Diagnostic performance of an ultra-sensitive RDT and a conventional RDT in malaria mass testing, treatment and tracking interventions in southern Ghana.

BACKGROUND: Application of numerous malaria control interventions has led to reduction in clinical malaria cases and deaths but also the realisation that asymptomatic parasite carriers play a key role in sustaining transmission. This study assessed the effectiveness of using the Ultra-sensitive NxTek eliminate RDT (uRDT) and conventional SD Bioline HRP2 RDT (cRDT) in diagnosing asymptomatic parasitaemia while measuring the impact of mass testing, treatment and tracking (MTTT) on the prevalence of asymptomatic malaria over a 1-year period in Ghana. METHODS: A total of 4000 targeted participants from two towns, Obom and Kofi Kwei, with their surrounding villages, were tested for asymptomatic malaria four times over the study period using uRDT (intervention) and the cRDT (control) respectively. Participants carrying malaria parasites were followed by home visit and phone calls for compliance to treatment, and filter paper blood blots collected from participants were used to determine true parasite carriage by PET-PCR. A mathematical model of the study site was developed and used to test the impact of test sensitivity and mass migration on the effect of MTTT. RESULTS: The start and end point sensitivities of the cRDT were 48.8% and 41.7% and those for the uRDT were 52.9% and 59.9% respectively. After a year of MTTTs, asymptomatic parasite prevalence, as determined by PCR, did not differ statistically in the control site (40.6% to 40.1%, P = 0.730) but decreased at the intervention site (55.9% to 46.4%, P < 0.0001). Parasite prevalence by RDT, however, indicated statistical reduction in the control site (25.3% to 22.3%, P = 0.017) and no change in the intervention site (35.1% to 36.0%, P = 0.614). The model predicted a mild effect of both diagnostic sensitivity and human movement in diminishing the impact of MTTT in the study sites. CONCLUSIONS: Asymptomatic parasite prevalence at the molecular level reduced significantly in the site where the uRDT was used but not where the cRDT was used. Overall, the uRDT exhibited higher sensitivity relative to the cRDT. Highly sensitive molecular techniques such as PET-PCR should be included in parasite prevalence estimation during MTTT exercises.

Screening of malaria infections in human blood samples with varying parasite densities and anaemic conditions using AI-Powered mid-infrared spectroscopy.

BACKGROUND: Effective testing for malaria, including the detection of infections at very low densities, is vital for the successful elimination of the disease. Unfortunately, existing methods are either inexpensive but poorly sensitive or sensitive but costly. Recent studies have shown that mid-infrared spectroscopy coupled with machine learning (MIRs-ML) has potential for rapidly detecting malaria infections but requires further evaluation on diverse samples representative of natural infections in endemic areas. The aim of this study was, therefore, to demonstrate a simple AI-powered, reagent-free, and user-friendly approach that uses mid-infrared spectra from dried blood spots to accurately detect malaria infections across varying parasite densities and anaemic conditions. METHODS: Plasmodium falciparum strains NF54 and FCR3 were cultured and mixed with blood from 70 malaria-free individuals to create various malaria parasitaemia and anaemic conditions. Blood dilutions produced three haematocrit ratios (50%, 25%, 12.5%) and five parasitaemia levels (6%, 0.1%, 0.002%, 0.00003%, 0%). Dried blood spots were prepared on Whatman™ filter papers and scanned using attenuated total reflection-Fourier Transform Infrared (ATR-FTIR) for machine-learning analysis. Three classifiers were trained on an 80%/20% split of 4655 spectra: (I) high contrast (6% parasitaemia vs. negative), (II) low contrast (0.00003% vs. negative) and (III) all concentrations (all positive levels vs. negative). The classifiers were validated with unseen datasets to detect malaria at various parasitaemia levels and anaemic conditions. Additionally, these classifiers were tested on samples from a population survey in malaria-endemic villages of southeastern Tanzania. RESULTS: The AI classifiers attained over 90% accuracy in detecting malaria infections as low as one parasite per microlitre of blood, a sensitivity unattainable by conventional RDTs and microscopy. These laboratory-developed classifiers seamlessly transitioned to field applicability, achieving over 80% accuracy in predicting natural P. falciparum infections in blood samples collected during the field survey. Crucially, the performance remained unaffected by various levels of anaemia, a common complication in malaria patients. CONCLUSION: These findings suggest that the AI-driven mid-infrared spectroscopy approach holds promise as a simplified, sensitive and cost-effective method for malaria screening, consistently performing well despite variations in parasite densities and anaemic conditions. The technique simply involves scanning dried blood spots with a desktop mid-infrared scanner and analysing the spectra using pre-trained AI classifiers, making it readily adaptable to field conditions in low-resource settings. In this study, the approach was successfully adapted to field use, effectively predicting natural malaria infections in blood samples from a population-level survey in Tanzania. With additional field trials and validation, this technique could significantly enhance malaria surveillance and contribute to accelerating malaria elimination efforts.

Preclinical evaluation of combined therapy with amiodarone and low-dose benznidazole in a mouse model of chronic Trypanosoma cruzi infection.

Chagasic chronic cardiomyopathy (CCC) is the primary clinical manifestation of Chagas disease (CD), caused by Trypanosoma cruzi. Current therapeutic options for CD are limited to benznidazole (Bz) and nifurtimox. Amiodarone (AMD) has emerged as most effective drug for treating the arrhythmic form of CCC. To address the effects of Bz and AMD we used a preclinical model of CCC. Female C57BL/6 mice were infected with T. cruzi and subjected to oral treatment for 30 consecutive days, either as monotherapy or in combination. AMD in monotherapy decreased the prolonged QTc interval, the incidence of atrioventricular conduction disorders and cardiac hypertrophy. However, AMD monotherapy did not impact parasitemia, parasite load, TNF concentration and production of reactive oxygen species (ROS) in cardiac tissue. Alike Bz therapy, the combination of Bz and AMD (Bz/AMD), improved cardiac electric abnormalities detected T. cruzi-infected mice such as decrease in heart rates, enlargement of PR and QTc intervals and increased incidence of atrioventricular block and sinus arrhythmia. Further, Bz/AMD therapy ameliorated the ventricular function and reduced parasite burden in the cardiac tissue and parasitemia to a degree comparable to Bz monotherapy. Importantly, Bz/AMD treatment efficiently reduced TNF concentration in the cardiac tissue and plasma and had beneficial effects on immunological abnormalities. Moreover, in the cardiac tissue Bz/AMD therapy reduced fibronectin and collagen deposition, mitochondrial damage and production of ROS, and improved sarcomeric and gap junction integrity. Our study underlines the potential of the Bz/AMD therapy, as we have shown that combination increased efficacy in the treatment of CCC.

Innate immune activation restricts priming and protective efficacy of the radiation-attenuated PfSPZ malaria vaccine.

A systems analysis was conducted to determine the potential molecular mechanisms underlying differential immunogenicity and protective efficacy results of a clinical trial of the radiation-attenuated whole-sporozoite PfSPZ vaccine in African infants. Innate immune activation and myeloid signatures at prevaccination baseline correlated with protection from P. falciparum parasitemia in placebo controls. These same signatures were associated with susceptibility to parasitemia among infants who received the highest and most protective PfSPZ vaccine dose. Machine learning identified spliceosome, proteosome, and resting DC signatures as prevaccination features predictive of protection after highest-dose PfSPZ vaccination, whereas baseline circumsporozoite protein-specific (CSP-specific) IgG predicted nonprotection. Prevaccination innate inflammatory and myeloid signatures were associated with higher sporozoite-specific IgG Ab response but undetectable PfSPZ-specific CD8+ T cell responses after vaccination. Consistent with these human data, innate stimulation in vivo conferred protection against infection by sporozoite injection in malaria-naive mice while diminishing the CD8+ T cell response to radiation-attenuated sporozoites. These data suggest a dichotomous role of innate stimulation for malaria protection and induction of protective immunity by whole-sporozoite malaria vaccines. The uncoupling of vaccine-induced protective immunity achieved by Abs from more protective CD8+ T cell responses suggests that PfSPZ vaccine efficacy in malaria-endemic settings may be constrained by opposing antigen presentation pathways.

A phase IIa, randomized, double-blind, safety, immunogenicity and efficacy trial of Plasmodium falciparum vaccine antigens merozoite surface protein 1 and RTS,S formulated with AS02 adjuvant in healthy, malaria-naïve adults.

BACKGROUND: To improve the efficacy of Plasmodium falciparum malaria vaccine RTS,S/AS02, we conducted a study in 2001 in healthy, malaria-naïve adults administered RTS,S/AS02 in combination with FMP1, a recombinant merozoite surface-protein-1, C-terminal 42kD fragment. METHODS: A double-blind Phase I/IIa study randomized N = 60 subjects 1:1:1:1 to one of four groups, N = 15/group, to evaluate safety, immunogenicity, and efficacy of intra-deltoid half-doses of RTS,S/AS02 and FMP1/AS02 administered in the contralateral (RTS,S + FMP1-separate) or same (RTS,S + FMP1-same) sites, or FMP1/AS02 alone (FMP1-alone), or RTS,S/AS02 alone (RTS,S-alone) on a 0-, 1-, 3-month schedule. Subjects receiving three doses of vaccine and non-immunized controls (N = 11) were infected with homologous P. falciparum 3D7 sporozoites by Controlled Human Malaria Infection (CHMI). RESULTS: Subjects in all vaccination groups experienced mostly mild or moderate local and general adverse events that resolved within eight days. Anti-circumsporozoite antibody levels were lower when FMP1 and RTS,S were co-administered at the same site (35.0 µg/mL: 95 % CI 20.3-63), versus separate arms (57.4 µg/mL: 95 % CI 32.3-102) or RTS,S alone (62.0 µg/mL: 95 % CI: 37.8-101.8). RTS,S-specific lymphoproliferative responses and ex vivo ELISpot CSP-specific interferon-gamma (IFN-γ) responses were indistinguishable among groups receiving RTS,S/AS02. There was no difference in antibody to FMP1 among groups receiving FMP1/AS02. After CHMI, groups immunized with a RTS,S-containing regimen had âˆ¼ 30 % sterile protection against parasitemia, and equivalent delays in time-to-parasitemia. The FMP1/AS02 alone group showed no sterile immunity or delay in parasitemia. CONCLUSION: Co-administration of RTS,S and FMP1/AS02 reduced anti-RTS,S antibody, but did not affect tolerability, cellular immunity, or efficacy in a stringent CHMI model. Absence of efficacy or delay of patency in the sporozoite challenge model in the FMP1/AS02 group did not rule out efficacy of FMP1/AS02 in an endemic population. However, a Phase IIb trial of FMP1/AS02 in children in malaria-endemic Kenya did not demonstrate efficacy against natural infection. CLINICALTRIALS: gov identifier: NCT01556945.

Oral infectivity through carnivorism in murine model of Trypanosoma cruzi infection.

Introduction: Oral transmission of T. cruzi is probably the most frequent transmission mechanism in wild animals. This observation led to the hypothesis that consuming raw or undercooked meat from animals infected with T. cruzi may be responsible for transmitting the infection. Therefore, the general objective of this study was to investigate host-pathogen interactions between the parasite and gastric mucosa and the role of meat consumption from infected animals in the oral transmission of T. cruzi. Methods: Cell infectivity assays were performed on AGS cells in the presence or absence of mucin, and the roles of pepsin and acidic pH were determined. Moreover, groups of five female Balb/c mice were fed with muscle tissue obtained from mice in the acute phase of infection by the clone H510 C8C3hvir of T. cruzi, and the infection of the fed mice was monitored by a parasitemia curve. Similarly, we assessed the infective capacity of T. cruzi trypomastigotes and amastigotes by infecting groups of five mice Balb/c females, which were infected orally using a nasogastric probe, and the infection was monitored by a parasitemia curve. Finally, different trypomastigote and amastigote inoculums were used to determine their infective capacities. Adhesion assays of T. cruzi proteins to AGS stomach cells were performed, and the adhered proteins were detected by western blotting using monoclonal or polyclonal antibodies and by LC-MS/MS and bioinformatics analysis. Results: Trypomastigote migration in the presence of mucin was reduced by approximately 30%, whereas in the presence of mucin and pepsin at pH 3.5, only a small proportion of parasites were able to migrate (∼6%). Similarly, the ability of TCTs to infect AGS cells in the presence of mucin is reduced by approximately 20%. In all cases, 60-100% of the animals were fed meat from mice infected in the acute phase or infected with trypomastigotes or amastigotes developed high parasitemia, and 80% died around day 40 post-infection. The adhesion assay showed that cruzipain is a molecule of trypomastigotes and amastigotes that binds to AGS cells. LC-MS/MS and bioinformatics analysis, also confirmed that transialidase, cysteine proteinases, and gp63 may be involved in TCTs attachment or invasion of human stomach cells because they can potentially interact with different proteins in the human stomach mucosa. In addition, several human gastric mucins have cysteine protease cleavage sites. Discussion: Then, under our experimental conditions, consuming meat from infected animals in the acute phase allows the T. cruzi infection. Similarly, trypomastigotes and amastigotes could infect mice when administered orally, whereas cysteinyl proteinases and trans-sialidase appear to be relevant molecules in this infective process.

Protective efficacy and safety of radiation-attenuated and chemo-attenuated Plasmodium Falciparum sporozoite vaccines against controlled and natural malaria infection: a systematic review and meta-analysis of randomized controlled trials.

BACKGROUND AND OBJECTIVE: Despite the significant burden of Plasmodium falciparum (Pf) malaria and the licensure of two vaccines for use in infants and young children that are partially effective in preventing clinical malaria caused by Pf, a highly effective vaccine against Pf infection is still lacking. Live attenuated vaccines using Pf sporozoites as the immunogen (PfSPZ Vaccines) hold promise for addressing this gap. Here we review the safety and efficacy of two of the most promising PfSPZ approaches: PfSPZ Vaccine (radiation attenuated PfSPZ) and PfSPZ-CVac (chemo-attenuated PfSPZ). METHODS: We conducted a systematic review and meta-analysis by searching PubMed, EMBASE, SCOPUS, CENTRAL, and WOS until 22nd December 2021. We included randomized controlled trials (RCTs) of these two vaccine approaches that measured protection against parasitaemia following controlled human malaria infection (CHMI) in malaria-naive and malaria-exposed adults or following exposure to naturally transmitted Pf malaria in African adults and children (primary outcome) and that also measured the incidence of solicited and unsolicited adverse events as indicators of safety and tolerability after vaccination (secondary outcome). We included randomized controlled trials (RCTs) that measured the detected parasitaemia after vaccination (primary outcome) and the incidence of various solicited and unsolicited adverse events (secondary outcome). The quality of the included RCTs using the Cochrane ROB 1 tool and the quality of evidence using the GRADE system were evaluated. We pooled dichotomous data using the risk ratio (RR) for development of parasitemia in vaccinees relative to controls as a measure of vaccine efficacy (VE), including the corresponding confidence interval (CI). This study was registered with PROSPERO (CRD42022308057). RESULTS: We included 19 RCTs. Pooled RR favoured PfSPZ Vaccine (RR: 0.65 with 95% CI [0.53, 0.79], P = 0.0001) and PfSPZ-table (RR: 0.42 with 95% CI [0.27, 0.67], P = 0.0002) for preventing parasitaemia, relative to normal saline placebo. Pooled RR showed no difference between PfSPZ Vaccine and the control in the occurrence of any solicited adverse event (RR: 1.00 with 95% CI [0.82, 1.23], P = 0.98), any local solicited adverse events (RR: 0.73 with 95% CI [0.49, 1.08], P = 0.11), any systemic solicited adverse events (RR: 0.94 with 95% CI [0.75, 1.17], P = 0.58), and any unsolicited adverse event (RR: 0.93 with 95% CI [0.78, 1.10], P = 0.37). CONCLUSION: PfSPZ and PfSPZ-CVacs showed comparable efficacy. Therefore, they can introduce a promising strategy for malaria prophylaxis, but more large-scale field trials are required to sustain efficacy and yield clinically applicable findings.

Multiple myeloma and Chagas disease: qPCR as a marker for preemptive antiparasitic therapy: a case reports series and review.

Multiple myeloma (MM) associated with Chagas disease is rarely described. This disease and its therapy suppress T cell and macrophage functions and increase regulatory T cell function, allowing the increase of parasitemia and the risk of Chagas Disease Reactivation (CDR). We aimed to analyze the role of conventional (cPCR) and quantitative Polymerase Chain Reaction (qPCR) for prospective monitoring of T. cruzi parasitemia, searching for markers of preemptive antiparasitic therapy in MM patients with Chagas disease. Moreover, we investigated the incidence and management of hematological diseases and CDR both inside and outside the transplant setting in the MEDLINE database. We found 293 studies and included 31 of them. Around 1.9-2.0% of patients with Chagas disease were reported in patients undergoing Stem Cell Transplantation. One case of CDR was described in eight cases of MM and Chagas disease. We monitored nine MM and Chagas disease patients, seven under Autologous Stem Cell Transplantation (ASCT), during 44.56±32.10 months (mean±SD) using parasitological methods, cPCR, and qPCR. From these patients, three had parasitemia. In the first, up to 256 par Eq/mL were detected, starting from 28 months after ASCT. The second patient dropped out and died soon after the detection of 161.0 par Eq/mL. The third patient had a positive blood culture. Benznidazole induced fast negativity in two cases; followed by notably lower levels in one of them. Increased T. cruzi parasitemia was related to the severity of the underlying disease. We recommend parasitemia monitoring by qPCR for early introduction of preemptive antiparasitic therapy to avoid CDR.

Schistosoma mansoni co-infection modulates Chagas disease development but does not impair the effect of benznidazole-based chemotherapy.

The adequate management of parasite co-infections represents a challenge that has not yet been overcome, especially considering that the pathological outcomes and responses to treatment are poorly understood. Thus, this study aimed to evaluate the impact of Schistosoma mansoni infection on the efficacy of benznidazole (BZN)-based chemotherapy in Trypanosoma cruzi co-infected mice. BALB/c mice were maintained uninfected or co-infected with S. mansoni and T. cruzi, and were untreated or treated with BZN. Body weight, mortality, parasitemia, cardiac parasitism, circulating cytokines (Th1/Th2/Th17); as well as heart, liver and intestine microstructure were analyzed. The parasitemia peak was five times higher and myocarditis was more severe in co-infected than T. cruzi-infected mice. After reaching peak, parasitemia was effectively controlled in co-infected animals. BZN successfully controlled parasitemia in both co-infected and T. cruzi-infected mice and improved body mass, cardiac parasitism, myocarditis and survival in co-infected mice. Co-infection dampened the typical cytokine response to either parasite, and BZN reduced anti-inflammatory cytokines in co-infected mice. Despite BZN normalizing splenomegaly and liver cellular infiltration, it exacerbated hepatomegaly in co-infected mice. Co-infection or BZN exerted no effect on hepatic granulomas, but increased pulmonary and intestinal granulomas. Marked granulomatous inflammation was identified in the small intestine of all schistosomiasis groups. Taken together, our findings indicate that BZN retains its therapeutic efficacy against T. cruzi infection even in the presence of S. mansoni co-infection, but with organ-specific repercussions, especially in the liver.

Changes in hypothalamic-pituitary-adrenal axis function, immunity, and glucose during acute Plasmodium relictum infection in house sparrows (Passer domesticus).

Hosts of the same species vary in physiological responses to the same parasite, and some groups of individuals can disproportionately affect disease dynamics; however, the underlying pathophysiology of host-parasite interactions is poorly understood in wildlife. We tested the hypothesis that the hypothalamic-pituitary-adrenal (HPA) axis mediates host resistance and tolerance to avian malaria during the acute phase of infection by evaluating whether individual variation in circulating glucocorticoids predicted resistance to avian malaria in a songbird. We experimentally inoculated wild-caught house sparrows (Passer domesticus) with naturally sourced Plasmodium relictum and quantified baseline and restraint-induced circulating corticosterone, negative feedback ability, cellular and humoral immune function, and baseline and restraint-induced glycemia, prior to and during acute malaria infection. During peak parasitemia, we also evaluated the expression of several liver cytokines that are established pathological hallmarks of malaria in mammals: two pro-inflammatory (IFN-γ and TNF-α) and two anti-inflammatory (IL-10 and TGF-ß). Although most of the host metrics we evaluated were not correlated with host resistance or tolerance to avian malaria, this experiment revealed novel relationships between malarial parasites and the avian immune system that further our understanding of the pathology of malaria infection in birds. Specifically, we found that: (1) TNF-α liver expression was positively correlated with parasitemia; (2) sparrows exhibited an anti-inflammatory profile during malaria infection; and (3) IFN-γ and circulating glucose were associated with several immune parameters, but only in infected sparrows. We also found that, during the acute phase of infection, sparrows increased the strength of corticosterone negative feedback at the level of the pituitary. In the context of our results, we discuss future methodological considerations and aspects of host physiology that may confer resistance to avian malaria, which can help inform conservation and rehabilitation strategies for avifauna at risk.

Assessment of biomarkers and clinical parameters as predictors of survival in patients with chagasic heart failure.

BACKGROUND: Chagas disease, endemic in Latin America and spreading globally due to emigration, has a significant health burden, particularly in relation to chagasic heart failure (HF). Chagasic cardiomyopathy (CCM) is characterized by chronic inflammatory myocardial disease. This study aimed to identify inflammatory parameters and biomarkers that could aid in the management of patients with chagasic HF. METHODS AND FINDINGS: A cohort study was conducted at a tertiary cardiology single-center over a mean follow-up period of 2.4 years. The study included patients with HF secondary to CCM enrolled between October 2013 and July 2017. Various clinical parameters, echocardiography findings, parasitemia status, brain natriuretic peptide (BNP) and troponin T (TnT) levels, and inflammatory biomarkers (IL-6, IL-10, IL-12p70, IL-17A, adiponectin, and IFN-γ) were assessed. The study encompassed a cohort of 103 patients, with a median age of 53 years and 70% being male. The left ventricular ejection fraction (LVEF) was 28%, with 40% of patients classified as NYHA II functional class. The median BNP level was 291 pg/ml. The observed mortality rate during the study period was 38.8%. Predictors of lower survival were identified as elevated levels of BNP, TnT, reduced LVEF, and increased adiponectin (thresholds: BNP > 309 pg/ml, TnT > 27.5 ng/ml, LVEF < 25.5%, adiponectin > 38 µg/mL). Notably, there was no evidence indicating a relationship between parasitemia and the inflammatory parameters with lower survival in these patients, including INF-γ, IL-6, IL-10, IL12-(p70), and IL17a. CONCLUSION: Despite the presence of a chronic inflammatory process, the evaluated inflammatory biomarkers in this cohort were not predictive of survival in patients with chagasic HF with reduced ejection fraction (HFrEF). However, reduced LVEF, elevated BNP, adiponectin levels, and troponin T were identified as predictors of lower survival in these patients.

[Investigation of the Efficacy of Cinnamaldehyde, Cannabidiol and Eravacycline in a Malaria Model]. / Sitma Modelinde Cinnamaldehyde, Cannabidiol ve Eravacycline'in Etkinliginin Arastirilmasi.

In this study, it was aimed to investigate the antimalarial activity of cinnamaldehyde (CIN) and cannabidiol (CBD) which have shown various biological activities such as potent antimicrobial activity and eravacycline (ERA), a new generation tetracycline derivative, in an in vivo malaria model. The cytotoxic activities of the active substances were determined by the MTT method against L929 mouse fibroblasts and their antimalarial activity were determined by the four-day test in an in vivo mouse model. In this study, five groups were formed: the CIN group, the CBD group, the ERA group, the chloroquine group (CQ) and the untreated group (TAG). 2.5 x 107 parasites/mL of P.berghei-infected erythrocyte suspension was administered IP to all mice. The determined doses of active substances were given to the mice by oral gavage in accordance with the four-day test and the parasitemia status in the mice was controlled for 21 days with smear preparations made from the blood taken from the tail end of the mice. The IC50 values, which express the cytotoxic activity values of the active substances were determined as 27.55 µg/mL, 16.40 µM and 48.82 µg/mL for CIN, CBD and ERA, respectively. The mean parasitemia rate in untreated mice was 33% on day nine and all mice died on day 11. On the ninth day, when compared with the TAG group, no parasites were observed in the CIN group, while the average parasitemia was 0.08% in the CBD group and 17.8% in the ERA group. Compared to the mice in the TAG group, the life expectancy of the other groups was prolonged by eight days in the CIN group, 12 days in the CBD group and eight days in the ERA group. It has been determined that all three active subtances tested in this study suppressed the development of Plasmodium parasites in an in vivo mouse model and prolonged the life span of the mice. It is thought that the strong antimalarial activity of CIN and CBD shown in the study and the possible positive effect of ERA on the clinical course can be improved by combining them with the existing and potential antimalarial molecules.

Anti-Malarial Activity of Allyl Isothiocyanate and N-acetyl-S-(N-allylthiocarbamoyl)-l-Cysteine.

SCOPE: Malaria remains one of the most important infectious diseases in the world. Allyl isothiocyanate (AITC) is a main ingredient of traditional spice Wasabia japonica, which is reported to have anti-bacterial and antiparasitic activities. However, there is no information on effects of AITC against malaria. The present study investigates the anti-malarial activity of dietary AITC in vivo and that of AITC metabolites in vitro. METHODS AND RESULTS: The ad libitum administration of 35, 175, or 350 µM AITC-containing drinking water to ICR mice significantly inhibit the parasitemia induced after infection with Plasmodium berghei. On the other hand, after single oral administration of AITC (20 mg kg-1 body weight), N-acetyl-S-(N-allylthiocarbamoyl)-l-cysteine (NAC-AITC) as one of the AITC metabolites displays a serum Cmax of 11.4 µM at a Tmax of 0.5 h, but AITC is not detected at any time point. Moreover, NAC-AITC shows anti-malarial activity against Plasmodium falciparum in vitro, and its 50% inhibitory concentration (IC50 ) against parasitemia is 12.6 µM. CONCLUSIONS: These results indicate that orally administered AITC is metabolized to NAC-AITC and exerts anti-malarial activity against malaria parasites in blood, suggesting that the consumption of AITC-containing food stuffs such as cruciferous plants may prevent malaria.

Haematological parameters and their correlation with the degree of malaria parasitaemia among outpatients attending a polyclinic.

BACKGROUND: Malaria is a parasitic disease caused by various species of the blood parasite Plasmodium; of all the parasitic diseases, malaria has the highest prevalence and mortality with an estimated 247 million cases and 619,000 deaths recorded worldwide as of 2021. Malaria causes febrile illness with several changes in blood cell parameters. Some of these changes include leucopenia, thrombocytopenia, and anaemia. If these changes could be correlated with the degree of parasitaemia, it can serve as a guide to physicians when treating malaria. This study was therefore aimed at correlating haematological parameters with levels of parasitaemia during malaria infection. METHODS: The study was a cross-sectional study involving 89 malaria positive patients. About 5 ml of blood was collected from each participant who gave his or her informed consent to partake in the study. A full blood count was performed on their samples to determine their haematological parameters using a haematology auto-analyzer. A parasite count was also performed via microscopy to determine the degree of parasitaemia. The data obtained from the study was entered into a database and statistically analysed using Statistical Package for Social Sciences (SPSS) version 23 and Microsoft Excel 2016. RESULTS: The study comprised of 89 participants out of which 35 were males and 54 were females with the mean age of 26.15 years. Secondary education participants were the highest with quaternary education the lowest. The highest parasite count recorded was 398,174 parasites/µl of blood, lowest count was 101 with the average being 32,942.32584. There was also a significant positive Pearson's correlation between total WBC and parasitaemia and with the WBC differentials, neutrophils, lymphocytes and monocytes had positive correlations while eosinophils and basophils had negative correlations. Furthermore, platelets, total RBC's, haemoglobin, MCH, MCHC and Hct all showed negative correlations. Linear regression also showed a linear relationship between parasite density and the various haematological parameters. CONCLUSION: The linear relationship (correlation) between WBC and MCH were the only significant ones at 95% and 99% confidence interval, respectively based on a two-tail t-test. Also, based on the regression analysis, the changes caused by WBC and PLT were the only significant changes at 95% confidence level in a two-tailed t-test.

The Toxoplasma gondii effector GRA83 modulates the host's innate immune response to regulate parasite infection.

Toxoplasma gondii's propensity to infect its host and cause disease is highly dependent on its ability to modulate host cell functions. One of the strategies the parasite uses to accomplish this is via the export of effector proteins from the secretory dense granules. Dense granule (GRA) proteins are known to play roles in nutrient acquisition, host cell cycle manipulation, and immune regulation. Here, we characterize a novel dense granule protein named GRA83, which localizes to the parasitophorous vacuole (PV) in tachyzoites and bradyzoites. Disruption of GRA83 results in increased virulence, weight loss, and parasitemia during the acute infection, as well as a marked increase in the cyst burden during the chronic infection. This increased parasitemia was associated with an accumulation of inflammatory infiltrates in tissues in both acute and chronic infections. Murine macrophages infected with ∆gra83 tachyzoites produced less interleukin-12 (IL-12) in vitro, which was confirmed with reduced IL-12 and interferon-gamma in vivo. This dysregulation of cytokines correlates with reduced nuclear translocation of the p65 subunit of the nuclear factor-κB (NF-κB) complex. While GRA15 similarly regulates NF-κB, infection with ∆gra83/∆gra15 parasites did not further reduce p65 translocation to the host cell nucleus, suggesting these GRAs function in converging pathways. We also used proximity labeling experiments to reveal candidate GRA83 interacting T. gondii-derived partners. Taken together, this work reveals a novel effector that stimulates the innate immune response, enabling the host to limit the parasite burden. Importance Toxoplasma gondii poses a significant public health concern as it is recognized as one of the leading foodborne pathogens in the United States. Infection with the parasite can cause congenital defects in neonates, life-threatening complications in immunosuppressed patients, and ocular disease. Specialized secretory organelles, including the dense granules, play an important role in the parasite's ability to efficiently invade and regulate components of the host's infection response machinery to limit parasite clearance and establish an acute infection. Toxoplasma's ability to avoid early clearance, while also successfully infecting the host long enough to establish a persistent chronic infection, is crucial in allowing for its transmission to a new host. While multiple GRAs directly modulate host signaling pathways, they do so in various ways highlighting the parasite's diverse arsenal of effectors that govern infection. Understanding how parasite-derived effectors harness host functions to evade defenses yet ensure a robust infection is important for understanding the complexity of the pathogen's tightly regulated infection. In this study, we characterize a novel secreted protein named GRA83 that stimulates the host cell's response to limit infection.

Use of PET/CT to detect myocardial inflammation and the risk of malignant arrhythmia in chronic Chagas disease.

BACKGROUND: Chagas heart disease (CHD) is characterized by progressive myocardial inflammation associated with myocardial fibrosis and segmental abnormalities that may lead to malignant ventricular arrhythmia and sudden cardiac death. This arrhythmia might be related to the persistence of parasitemia or inflammation in the myocardium in late-stage CHD. Positron emission tomography/computed tomography (PET/CT) has been used to detect myocardial inflammation in non-ischemic cardiomyopathies, such as sarcoidosis, and might be useful for risk prediction in patients with CHD. METHODS AND RESULTS: Twenty-four outpatients with chronic CHD were enrolled in this prospective cross-sectional study between May 2019 and March 2022. The patients were divided into two groups: those with sustained ventricular tachycardia and/or aborted sudden cardiac death who required implantable cardioverter-defibrillators, and those with the same stages of CHD and no complex ventricular arrhythmia. Patients underwent 18F-fluorodeoxyglucose (18F-FDG) and 68Ga-DOTATOC PET/CT, and blood samples were collected for qualitative parasite assessment by polymerase chain reaction. Although similar proportions of patients with and without complex ventricular arrhythmia showed 18F-FDG and 68Ga-DOTATOC uptake, 68Ga-DOTATOC corrected SUVmax was higher in patients with complex arrhythmia (3.4 vs 1.7; P = .046), suggesting that inflammation could be associated with the presence of malignant arrhythmia in the late stages of CHD. We also detected Trypanosoma cruzi in both groups, with a nonsignificant trend of increased parasitemia in the group with malignant arrhythmia (66.7% vs 33.3%). CONCLUSION: 18F-FDG and 68Ga-DOTATOC uptake on PET/CT may be useful for the detection of myocardial inflammation in patients with Chagas cardiomyopathy, and 68Ga-DOTATOC uptake may be associated with the presence of malignant arrhythmia, with potential therapeutic implications.

Design and synthesis of new 1,2,3-triazoles derived from eugenol and analogues with in vitro and in vivo activity against Trypanosoma cruzi.

Chagas disease (CD) is a neglected tropical disease endemic in 21 countries and affects about 8 million people around the world. The pharmacotherapy for this disease is limited to two drugs (Benznidazole and Nifurtimox) and both are associated with important limitations, as low cure rate in the chronic phase of the disease, high toxicity and increasing resistance by Trypanosoma cruzi. Recently, we reported a bioactive 1,2,3-triazole (compound 35) active in vitro (IC50 42.8 µM) and in vivo (100 mg/kg) against T. cruzi Y strains and preliminary in silico studies suggested the cysteine protease cruzain as a possible target. Considering these initial findings, we describe here the design and synthesis of new 1,2,3-triazoles derivatives of our hit compound (35). The triazoles were initially evaluated against healthy cells derived from neonatal rat cardiomyoblasts (H9c2 cells) to determine their cytotoxicity and against epimastigotes forms of T. cruzi Y strain. The most active triazoles were compounds 26 (IC50 19.7 µM) and 27 (IC50 7.3 µM), while benznidazole was active at 21.6 µM. Derivative 27 showed an interesting selectivity index considering healthy H9c2 cells (>77). Promising activities against trypomastigotes forms of the parasite were also observed for triazoles 26 (IC50 20.74 µM) and 27 (IC50 8.41 µM), mainly 27 which showed activity once again higher than that observed for benznidazole (IC50 12.72 µM). While docking results suggested cruzain as a potential target for these compounds, no significant enzyme inhibition was observed in vitro, indicating that their trypanocidal activity is related to another mode of action. Considering the promising in vitro results of triazoles 26 and 27, the in vivo toxicity was initially verified based on the evaluation of behavioral and physiological parameters, mortality, effect in body weight gain, and through the measurement of AST/ALT enzymes, which are markers of liver toxicity. All these evaluations pointed to a good tolerability of the animals, especially considering triazole 27. A reduction in parasitemia was observed among animals treated with triazole 27, but not among those treated with derivative 26. Regarding the dosage, derivative 27 (100 mg/kg) was the most active sample against T. cruzi infection, showing a 99.4% reduction in parasitemia peak. Triazole 27 at a dosage of 100 mg/kg influenced the humoral immune response and reduced myocarditis in the animals, bringing antibody levels closer to those observed among healthy mice. Altogether, our results indicate compound 27 as a new lead for the development of drug candidates to treat Chagas disease.

Vitamin C-rich juice co-administration with artemether-lumefantrine ameliorates oxido-inflammatory responses in Plasmodium berghei-infected mice.

This study investigated the effects of co-administration of a commercial juice rich in vitamin C (Vit C) on the antimalarial efficacy of artemether-lumefantrine (AL) in Plasmodium berghei-infected mice. Fifty Balb/c mice were infected with Plasmodium berghei NK65 strain from a donor mouse. Parasitemia was established after 72 h. Animals were grouped into 6 (n = 10) and treated daily for 3 days with normal saline, chloroquine, artemether-lumefantrine (AL), AL plus 50% commercial juice (CJ), and AL plus 50% Vit C supplementation in drinks ad libitum, respectively. Body weight, parasitemia levels, and mean survival time were determined. Tumor necrosis factor-alpha (TNF-α), interleukin 6 (IL-6), nitrite, malondialdehyde, reduced glutathione (GSH), catalase, and superoxide dismutase (SOD) were determined in the serum and liver tissues. Spleen histopathological changes were determined by H&E staining. Parasitemia was cleared by administration of AL and was not affected by Vit C and CJ supplementation. Vit C significantly prevented body weight reduction in AL-treated mice. CJ and Vit C supplementation to AL-treated mice significantly improved survival proportion compared with AL alone animals. Vit C and CJ supplementation significantly improved reduction of TNF-α, IL-6, and malondialdehyde, and increased GSH, CAT, and SOD in AL-treated mice. Spleen cell degeneration and presence of malaria pigment were reduced in AL-treated animals. The results suggest that ad libitum co-administration of commercial juice and vitamin C with artemether-lumefantrine does not impair its antimalarial efficacy but rather improved antioxidant and anti-inflammatory effects in mice.

Cysteine Residues in Region 6 of the Plasmodium yoelii Erythrocyte-Binding-like Ligand That Are Related to Its Localization and the Course of Infection.

Plasmodium malaria parasites use erythrocyte-binding-like (EBL) ligands to invade erythrocytes in their vertebrate host. EBLs are released from micronemes, which are secretory organelles located at the merozoite apical end and bind to erythrocyte surface receptors. Because of their essential nature, EBLs have been studied as vaccine candidates, such as the Plasmodium vivax Duffy binding protein. Previously, we showed through using the rodent malaria parasite Plasmodium yoelii that a single amino acid substitution within the EBL C-terminal Cys-rich domain (region 6) caused mislocalization of this molecule and resulted in alteration of the infection course and virulence between the non-lethal 17X and lethal 17XL strains. In the present study, we generated a panel of transgenic P. yoelii lines in which seven of the eight conserved Cys residues in EBL region 6 were independently substituted to Ala residues to observe the consequence of these substitutions with respect to EBL localization, the infection course, and virulence. Five out of seven transgenic lines showed EBL mislocalizations and higher parasitemias. Among them, three showed increased virulence, whereas the other two did not kill the infected mice. The remaining two transgenic lines showed low parasitemias similar to their parental 17X strain, and their EBL localizations did not change. The results indicate the importance of Cys residues in EBL region 6 for EBL localization, parasite infection course, and virulence and suggest an association between EBL localization and the parasite infection course.

Neither the African-Centric S47 Nor P72 Variant of TP53 Is Associated With Reduced Risk of Febrile Malaria in a Malian Cohort Study.

BACKGROUND: TP53 has been shown to play a role in inflammatory processes, including malaria. We previously found that p53 attenuates parasite-induced inflammation and predicts clinical protection to Plasmodium falciparum infection in Malian children. Here, we investigated whether p53 codon 47 and 72 polymorphisms are associated with differential risk of P. falciparum infection and uncomplicated malaria in a prospective cohort study of malaria immunity. METHODS: p53 codon 47 and 72 polymorphisms were determined by sequencing TP53 exon 4 in 631 Malian children and adults enrolled in the Kalifabougou cohort study. The effects of these polymorphisms on the prospective risk of febrile malaria, incident parasitemia, and time to fever after incident parasitemia over 6 months of intense malaria transmission were assessed using Cox proportional hazards models. RESULTS: Confounders of malaria risk, including age and hemoglobin S or C, were similar between individuals with or without p53 S47 and R72 polymorphisms. Relative to their respective common variants, neither S47 nor R72 was associated with differences in prospective risk of febrile malaria, incident parasitemia, or febrile malaria after parasitemia. CONCLUSIONS: These findings indicate that p53 codon 47 and 72 polymorphisms are not associated with protection against incident P. falciparum parasitemia or uncomplicated febrile malaria.