Parasite Strains Research Articles

First experimental observation on biology of the avian malaria parasite Plasmodium (Novyella) homonucleophilum (lineage pSW2), with remarks on virulence and distribution

Species of subgenus Novyella remain most fragmentarily studied amongst avian malaria agents. Transmission of the recently described Plasmodium (Novyella) homonucleophilum (lineage pSW2) occurs broadly in the Old World, including Europe, however biology of this pathogen remains insufficiently investigated. This study provided the first data on the development of P. homonucleophilum in the experimentally infected Eurasian siskins Spinus spinus exposed by inoculation of infected blood. The parasite strain was isolated from a naturally infected song thrush Turdus philomelos, multiplied in vivo, and inoculated to six Eurasian siskins. The same number of birds were used as negative controls. All exposed birds were susceptible, and the controls remained uninfected during the entire study (172 days). Prepatent period was 8–12 days post exposure (dpe). Maximum parasitaemia reached 50–90 % of infected erythrocytes between 20 and 44 dpe. Then, parasitaemia decreased but remained relatively high during the entire observation. Three of six exposed birds died, indicating high virulence of this infection. The parasitaemia increase coincided with a decline of haematocrit value, indicating anaemia. Polychromasia was evident in all infected birds but not in controls. Body mass of exposed birds increased, coinciding with increased food intake. The latter probably is an adaptation to compensate energy loss of hosts due to the long-lasting parasitism. Exo-erythrocytic stages were not found, suggesting that long-lasting parasitaemia was entirely due to erythrocytic merogony. The lineage pSW2 has been reported broadly in the Old World and is likely a generalist infection. Neglected avian Novyella malaria parasites are worth more attention of researchers due to their cosmopolitan distribution and high virulence.

Evaluating the performance of Plasmodium falciparum genetic metrics for inferring National Malaria Control Programme reported incidence in Senegal

BackgroundGenetic surveillance of the Plasmodium falciparum parasite shows great promise for helping National Malaria Control Programmes (NMCPs) assess parasite transmission. Genetic metrics such as the frequency of polygenomic (multiple strain) infections, genetic clones, and the complexity of infection (COI, number of strains per infection) are correlated with transmission intensity. However, despite these correlations, it is unclear whether genetic metrics alone are sufficient to estimate clinical incidence.MethodsThis study examined parasites from 3147 clinical infections sampled between the years 2012–2020 through passive case detection (PCD) across 16 clinic sites spread throughout Senegal. Samples were genotyped with a 24 single nucleotide polymorphism (SNP) molecular barcode that detects parasite strains, distinguishes polygenomic (multiple strain) from monogenomic (single strain) infections, and identifies clonal infections. To determine whether genetic signals can predict incidence, a series of Poisson generalized linear mixed-effects models were constructed to predict the incidence level at each clinical site from a set of genetic metrics designed to measure parasite clonality, superinfection, and co-transmission rates.ResultsModel-predicted incidence was compared with the reported standard incidence data determined by the NMCP for each clinic and found that parasite genetic metrics generally correlated with reported incidence, with departures from expected values at very low annual incidence (< 10/1000/annual [‰]).ConclusionsWhen transmission is greater than 10 cases per 1000 annual parasite incidence (annual incidence > 10‰), parasite genetics can be used to accurately infer incidence and is consistent with superinfection-based hypotheses of malaria transmission. When transmission was < 10‰, many of the correlations between parasite genetics and incidence were reversed, which may reflect the disproportionate impact of importation and focal transmission on parasite genetics when local transmission levels are low.

Stimulation of microvesicle secretion in Trichomonas vaginalis

Trichomonas vaginalis is an extracellular flagellate protozoan and the etiological agent of human trichomoniasis, a sexually transmitted infection (STI) with a high incidence. Several reports have shown that this protozoan releases microvesicles into the culture medium, which show high potential in modulating cell-to-cell communication and the host response to infections. However, the biogenesis of these vesicles has not been analyzed in detail. In the present study, high-resolution ion scanning microscopy (SEM) and transmission electron microscopy (TEM) were used to analyze the surface of control cells and cells incubated in the presence of Ca2+ alone or with A 23187 calcium ionophore. Two different strains of T. vaginalis were analyzed. Most control cells displayed relatively smooth surfaces, whereas cells incubated with Ca2+ had many surface projections of variable shape and size (from 40 nm to around 1 μm). Quantitative analyses were performed directly in the scanning electron microscope and showed a significant increase in the number of cells with surface projections after incubation in the presence of calcium. TEM showed that treated cells presented several cytoplasmic multivesicular structures, suggesting membrane fusion and exosomes in the extracellular medium. The amount and size of the released vesicles were quantitatively analyzed using light scattering and TEM on negatively stained samples.The observations show that incubation of both parasite strains in the presence of Ca2+ significantly increased the release of microvesicles into the extracellular medium in a time-dependent process. Sequential incubation in the presence of Ca2+ and the calcium ionophore A23187 increases the presence of vesicles on the parasite surface only at a short incubation time (5 min). Transmission electron microscopy showed that at least part of the vesicles are originated from cytoplasmic multivesicular structures. This information contributes to a better understanding of the biogenesis of extracellular vesicles secreted by T. vaginalis.

Antigenic strain diversity predicts different biogeographic patterns of maintenance and decline of antimalarial drug resistance.

The establishment and spread of antimalarial drug resistance vary drastically across different biogeographic regions. Though most infections occur in sub-Saharan Africa, resistant strains often emerge in low-transmission regions. Existing models on resistance evolution lack consensus on the relationship between transmission intensity and drug resistance, possibly due to overlooking the feedback between antigenic diversity, host immunity, and selection for resistance. To address this, we developed a novel compartmental model that tracks sensitive and resistant parasite strains, as well as the host dynamics of generalized and antigen-specific immunity. Our results show a negative correlation between parasite prevalence and resistance frequency, regardless of resistance cost or efficacy. Validation using chloroquine-resistant marker data supports this trend. Post discontinuation of drugs, resistance remains high in low-diversity, low-transmission regions, while it steadily decreases in high-diversity, high-transmission regions. Our study underscores the critical role of malaria strain diversity in the biogeographic patterns of resistance evolution.

Antigenic strain diversity predicts different biogeographic patterns of maintenance and decline of antimalarial drug resistance

The establishment and spread of antimalarial drug resistance vary drastically across different biogeographic regions. Though most infections occur in sub-Saharan Africa, resistant strains often emerge in low-transmission regions. Existing models on resistance evolution lack consensus on the relationship between transmission intensity and drug resistance, possibly due to overlooking the feedback between antigenic diversity, host immunity, and selection for resistance. To address this, we developed a novel compartmental model that tracks sensitive and resistant parasite strains, as well as the host dynamics of generalized and antigen-specific immunity. Our results show a negative correlation between parasite prevalence and resistance frequency, regardless of resistance cost or efficacy. Validation using chloroquine-resistant marker data supports this trend. Post discontinuation of drugs, resistance remains high in low-diversity, low-transmission regions, while it steadily decreases in high-diversity, high-transmission regions. Our study underscores the critical role of malaria strain diversity in the biogeographic patterns of resistance evolution.

A biological invasion modifies the dynamics of a host-parasite arms race.

By imposing novel selection pressures on both participants, biological invasions can modify evolutionary 'arms races' between hosts and parasites. A spatially replicated cross-infection experiment reveals strong spatial divergence in the ability of lungworms (Rhabdias pseudosphaerocephala) to infect invasive cane toads (Rhinella marina) in Australia. In areas colonized for longer than 20 years, toads are more resistant to infection by local strains of parasites than by allopatric strains. The situation reverses at the invasion front, where super-infective parasites have evolved. Invasion-induced shifts in genetic diversity and selective pressures may explain why hosts gain advantage over parasites in long-colonized areas, whereas parasites gain advantage at the invasion front.

Enhanced Antimalarial and Antisequestration Activity of Methoxybenzenesulfonate-Modified Biopolymers and Nanoparticles for Tackling Severe Malaria.

Severe malaria is a life-threatening condition that is associated with a high mortality. Severe Plasmodium falciparum infections are mediated primarily by high parasitemia and binding of infected red blood cells (iRBCs) to the blood vessel endothelial layer, a process known as sequestration. Here, we show that including the 5-amino-2-methoxybenzenesulfonate (AMBS) chemical modification in soluble biopolymers (polyglutamic acid and heparin) and poly(acrylic acid)-exposing nanoparticles serves as a universal tool to introduce a potent parasite invasion inhibitory function in these materials. Importantly, the modification did not add or eliminated (for heparin) undesired anticoagulation activity. The materials protected RBCs from invasion by various parasite strains, employing both major entry pathways. Two further P. falciparum strains, which either expose ligands for chondroitin sulfate A (CSA) or intercellular adhesion molecule 1 (ICAM-1) on iRBCs, were tested in antisequestration assays due to their relevance in placental and cerebral malaria, respectively. Antisequestration activity was found to be more efficacious with nanoparticles vs gold-standard soluble biopolymers (CSA and heparin) against both strains, when tested on receptor-coated dishes. The nanoparticles also efficiently inhibited and reversed the sequestration of iRBCs on endothelial cells. First, the materials described herein have the potential to reduce the parasite burden by acting at the key multiplication stage of reinvasion. Second, the antisequestration ability could help remove iRBCs from the blood vessel endothelium, which could otherwise cause vessel obstruction, which in turn can lead to multiple organ failure in severe malaria infections. This approach represents a further step toward creation of adjunctive therapies for this devastating condition to reduce morbidity and mortality.

Assessment of the Performance of Lactate Dehydrogenase-Based Rapid Diagnostic Test for Malaria in Djibouti in 2022–2023

Until 2020, Djiboutian health authorities relied on histidine-rich protein-2 (HRP2)-based rapid diagnostic tests (RDTs) to establish the diagnosis of Plasmodium falciparum. The rapid spread of P. falciparum histidine-rich protein-2 and -3 (pfhrp2/3) gene-deleted parasite strains in Djibouti has led the authorities to switch from HRP2-based RDTs to lactate dehydrogenase (LDH)-based RDTs targeting the plasmodial lactate dehydrogenase (pLDH) specific for P. falciparum and P. vivax (RapiGEN BIOCREDIT Malaria Ag Pf/Pv pLDH/pLDH) in 2021. This study was conducted with the primary objective of evaluating the diagnostic performance of this alternative RDT. Operational constraints related, in particular, to the implementation of this RDT during the COVID-19 pandemic were also considered. The performance of BIOCREDIT Malaria Ag Pf/Pv (pLDH/pLDH) RDT was also compared to our previously published data on the performance of two HRP2-based RDTs deployed in Djibouti in 2018–2020. The diagnosis of 350 febrile patients with suspected malaria in Djibouti city was established using two batches of RapiGEN BIOCREDIT Malaria Ag Pf/Pv (pLDH/pLDH) RDT over a two-year period (2022 and 2023) and confirmed by real-time quantitative polymerase chain reaction. The sensitivity and specificity for the detection of P. falciparum were 88.2% and 100%, respectively. For P. vivax, the sensitivity was 86.7% and the specificity was 100%. Re-training and closer supervision of the technicians between 2022 and 2023 have led to an increased sensitivity to detect P. falciparum (69.8% in 2022 versus 88.2% in 2023; p &lt; 0.01). The receiver operating characteristic curve analysis highlighted a better performance in the diagnosis of P. falciparum with pLDH-based RDTs compared with previous HRP2-based RDTs. In Djibouti, where pfhrp2-deleted strains are rapidly gaining ground, LDH-based RDTs seem to be more suitable for diagnosing P. falciparum than HRP2-based RDTs. Awareness-raising and training for technical staff have also been beneficial.

Bioassay-guided isolation of leishmanicidal cucurbitacins from Momordica charantia.

Leishmaniasis, a neglected tropical parasitic disease, is regarded as a major public health problem worldwide. The first-line drugs for leishmaniasis suffer from limitations related to toxicity and the development of resistance in certain parasitic strains. Therefore, the discovery of alternative treatments for leishmaniasis is imperative, and natural products represent a valuable source of potential therapeutic agents. The present study aimed at finding new potential antileishmanial agents from the aerial parts of the medicinal plant Momordica charantia. This study was based on bioassay-guided fractionation of the M. charantia extract against promastigotes and amastigotes of Leishmania (Leishmania) amazonensis. The cytotoxicity of the extract, fractions, and isolated compounds were evaluated against peritoneal murine macrophages by employing the MTT assay for assessing cell metabolic activity. Antileishmanial assay-guided fractionation of the M. charantia extract led to the bioactive cucurbitacin-enriched fraction and the isolation of four bioactive cucurbitacin-type triterpenoids, which exhibited significant antileishmanial activity, with IC50 values between 2.11 and 3.25μg.mL-1 against promastigote and amastigote forms, low toxicity and selectivity indexes ranging from 8.5 to 17.2. Our findings demonstrate that the fractions and cucurbitacin-type triterpenoids obtained from the aerial parts of M. charantia are promising natural leishmanicidal candidates.

Antimalarial Drug Resistance in Africa: Current Status and Future Prospects

Malaria remains a significant public health concern in Africa, with millions of cases reported annually. However, the effectiveness of antimalarial drugs, a cornerstone of malaria control efforts, is threatened by the emergence and spread of drug-resistant strains of the Plasmodium parasite. This article provides a comprehensive overview of the current status of antimalarial drug resistance in Africa and explores prospects for combatting this critical issue. In recent years, various regions across the continent have witnessed the rise of drug-resistant malaria strains, posing formidable challenges to malaria treatment and control programs. We delve into the underlying genetic and molecular mechanisms driving drug resistance, shedding light on the role of mutations and the evolutionary forces at play. Despite these challenges, ongoing research and global collaborations offer hope for the future. Combination therapies, innovative drug development, and vector control measures are discussed as strategies to mitigate drug resistance. We also explore the potential of genomics and molecular studies in understanding and countering resistance. However, significant barriers persist, including limited funding, resource disparities, and access to healthcare in remote areas. Socioeconomic factors further complicate the landscape of malaria control. In conclusion, this article underscores the urgency of addressing antimalarial drug resistance in Africa as a public health priority. By staying informed about the current status, supporting research initiatives, and promoting effective policies and interventions, we can strive for a future where malaria treatment remains effective and accessible to all, ultimately working toward the goal of malaria eradication on the African continent.

"Seasoning" antimalarial drugs' action: chloroquine bile salts as novel triple-stage antiplasmodial hits.

Malaria is one of the "big three" global infectious diseases, having caused above two hundred million cases and over half a million deaths in 2020. The continuous demand for new treatment options prioritizes the cost-effective development of new chemical entities with multi-stage antiplasmodial activity, for higher efficacy and lower propensity to elicit drug-resistant parasite strains. Following up on our long-term research towards the rescue of classical antimalarial aminoquinolines like chloroquine and primaquine, we have developed new organic salts by acid-base pairing of those drugs with natural bile acids. These antimalarial drug-derived bile salts were screened in vitro against the hepatic, blood and gametocyte stages of Plasmodium parasites, unveiling chloroquine bile salts as unprecedented triple-stage antiplasmodial hits. These findings pave a new pathway for drug rescuing, even beyond anti-malarial and other anti-infective drugs.

Increased sensitivity of malaria parasites to common antimalaria drugs after the introduction of artemether-lumefantrine: Implication of policy change and implementation of more effective drugs in fight against malaria.

Single nucleotide polymorphisms (SNPs) in the Plasmodium falciparum multi-drug resistance protein 1 (Pfmrp1) gene have previously been reported to confer resistance to Artemisinin-based Combination Therapies (ACTs) in Southeast Asia. A total of 300 samples collected from six sites between 2008 and 2019 under an ongoing malaria drug sensitivity patterns in Kenya study were evaluated for the presence of SNPs at Pfmrp1 gene codons: H191Y, S437A, I876V, and F1390I using the Agena MassARRAY® platform. Each isolate was further tested against artemisinin (ART), lumefantrine (LU), amodiaquine (AQ), mefloquine (MQ), quinine (QN), and chloroquine (CQ) using malaria the SYBR Green I-based method to determine their in vitro drug sensitivity. Of the samples genotyped, polymorphism at Pfmrp1 codon I876V was the most frequent, with 59.3% (163/275) mutants, followed by F1390I, 7.2% (20/278), H191Y, 4.0% (6/151), and S437A, 3.3% (9/274). A significant decrease in median 50% inhibition concentrations (IC50s) and interquartile range (IQR) was noted; AQ from 2.996 ng/ml [IQR = 2.604-4.747, n = 51] in 2008 to 1.495 ng/ml [IQR = 0.7134-3.318, n = 40] (P<0.001) in 2019, QN from 59.64 ng/ml [IQR = 29.88-80.89, n = 51] in 2008 to 18.10 ng/ml [IQR = 11.81-26.92, n = 42] (P<0.001) in 2019, CQ from 35.19 ng/ml [IQR = 16.99-71.20, n = 30] in 2008 to 6.699 ng/ml [IQR = 4.976-9.875, n = 37] (P<0.001) in 2019, and ART from 2.680 ng/ml [IQR = 1.608-4.857, n = 57] in 2008 to 2.105 ng/ml [IQR = 1.266-3.267, n = 47] (P = 0.0012) in 2019, implying increasing parasite sensitivity to the drugs over time. However, no significant variations were observed in LU (P = 0.2692) and MQ (P = 0.0939) respectively, suggesting stable parasite responses over time. There was no statistical significance between the mutation at 876 and parasite sensitivity to selected antimalarials tested, suggesting stable sensitivity for the parasites with 876V mutations. These findings show that Kenyan parasite strains are still sensitive to AQ, QN, CQ, ART, LU, and MQ. Despite the presence of Pfmrp1 mutations in parasites among the population.

A tale of two parasites: Responses of honey bees infected with Nosema ceranae and Lotmaria passim

Nosema ceranae and Lotmaria passim are two commonly encountered digestive tract parasites of the honey bee that have been associated with colony losses in Canada, the United States, and Europe. Though honey bees can be co-infected with these parasites, we still lack basic information regarding how they impact bee health at the individual and colony level. Using locally-isolated parasite strains, we investigated the effect of single and co-infections of these parasites on individual honey bee survival, and their responsiveness to sucrose. Results showed that a single N. ceranae infection is more virulent than both single L. passim infections and co-infections. Honey bees singly infected with N. ceranae reached < 50% survival eight days earlier than those inoculated with L. passim alone, and four days earlier than those inoculated with both parasites. Honey bees infected with either one, or both, parasites had increased responsiveness to sucrose compared to uninfected bees, which could correspond to higher levels of hunger and increased energetic stress. Together, these findings suggest that N. ceranae and L. passim pose threats to bee health, and that the beekeeping industry should monitor for both parasites in an effort correlate pathogen status with changes in colony-level productivity and survival.

Susceptibility of Toxoplasma gondii to autophagy in human cells relies on multiple interacting parasite loci.

Autophagy is a process used by cells to recycle organelles and macromolecules and to eliminate intracellular pathogens. Previous studies have shown that some stains of Toxoplasma gondii are resistant to autophagy-dependent growth restriction, while others are highly susceptible. Although it is known that autophagy-mediated control requires activation by interferon gamma, the basis for why parasite strains differ in their susceptibility is unknown. Our findings indicate that susceptibility involves at least five unlinked parasite genes on different chromosomes, including several secretory proteins targeted to the parasite-containing vacuole and exposed to the host cell cytosol. Our findings reveal that susceptibility to autophagy-mediated growth restriction relies on differential recognition of parasite proteins exposed at the host-pathogen interface, thus identifying a new mechanism for cell-autonomous control of intracellular pathogens.

Blood-stage antimalarial activity, favourable metabolic stability and in vivo toxicity of novel piperazine linked 7-chloroquinoline-triazole conjugates

The persistence of drug resistance poses a significant obstacle to the advancement of efficacious malaria treatments. The remarkable efficacy displayed by 1,2,3-triazole-based compounds against Plasmodium falciparum highlights the potential of triazole conjugates, with diverse pharmacologically active structures, as potential antimalarial agents. We aimed to synthesize 7-dichloroquinoline-triazole conjugates and their structure-activity relationship (SAR) derivatives to investigate their anti-plasmodial activity. Among them, QP11, featuring a m-NO2 substitution, demonstrated efficacy against both chloroquine-sensitive and -resistant parasite strains. QP11 selectively inhibited FP2, a cysteine protease involved in hemoglobin degradation, and showed synergistic effects when combined with chloroquine. Additionally, QP11 hindered hemoglobin degradation and hemozoin formation within the parasite. Metabolic stability studies indicated high stability of QP11, making it a promising antimalarial candidate. In vivo evaluation using a murine malaria model demonstrated QP11's efficacy in eradicating parasite growth without neurotoxicity, presenting it as a promising compound for novel antimalarial development.

Antiplasmodial Activity of β-Ionone and the Effect of the Compound on Amelioration of Anaemia and Oxidative Organ Damage in Mice Infected with Plasmodium berghei.

Owing to evolution of parasite strains that are resistant to existing antimalarial drugs, research for novel antimalarial medicines is progressing on numerous fronts. Herein, we evaluated the in vivo anti-Plasmodium berghei activity of β-ionone including its ameliorative potential towards P. berghei-associated anaemia and oxidative organ damage. Mice were infected with chloroquine-sensitive strain of P. berghei and then treated with β-ionone at doses of 10 and 20mg/kg body weight (BW) for seven days. The parasitemia, packed cell volume and redox sensitive biomarkers in the liver, brain and spleen were estimated. Our result showed that β-ionone, in a dose-dependent fashion, significantly (p < 0.05) repressed the multiplication of P. berghei. More so, the compound, at doses of 10 and 20mg/kg BW, significantly (p < 0.05) mitigated anaemia and organ damage induced by P. berghei. Overall, the findings demonstrated that β-ionone has antiplasmodial actions and plays a mitigative role against P. berghei-induced anaemia and oxidative organ damage.

Naturally Acquired Transmission-Blocking Immunity Against Different Strains of Plasmodium vivax in a Malaria-Endemic Area in Thailand.

Human immunity triggered by natural malaria infections impedes parasite transmission from humans to mosquitoes, leading to interest in transmission-blocking vaccines. However, immunity characteristics, especially strain specificity, remain largely unexplored. We investigated naturally acquired transmission-blocking immunity (TBI) against Plasmodium vivax, a major malaria parasite. Using the direct membrane-feeding assay, we assessed TBI in plasma samples and examined the role of antibodies by removing immunoglobulins through protein G/L adsorption before mosquito feeding. Strain specificity was evaluated by conducting a direct membrane-feeding assay with plasma exchange. Blood samples from 47 patients with P vivax were evaluated, with 37 plasma samples successfully infecting mosquitoes. Among these, 26 showed inhibition before immunoglobulin depletion. Despite substantial immunoglobulin removal, 4 samples still exhibited notable inhibition, while 22 had reduced blocking activity. Testing against heterologous strains revealed some plasma samples with broad TBI and others with strain-specific TBI. Our findings indicate that naturally acquired TBI is mainly mediated by antibodies, with possible contributions from other serum factors. The transmission-blocking activity of plasma samples varied by the tested parasite strain, suggesting single polymorphic or multiple targets for naturally acquired TBI. These observations improve understanding of immunity against P vivax and hold implications for transmission-blocking vaccine development.

Medicinal plants with antileishmanial activity on parasites responsible for new-world cutaneous leishmaniasis. A systematic review 2018-2022

Context: Cutaneous leishmaniasis is a disease of public health importance; treatment is based on the use of pentavalent antimonials with high toxicity and low efficacy; therefore, it´s necessary to search for therapeutic alternatives derived from natural products, based on the study of medicinal plants as a source of molecules with highly effective leishmanicidal potential. Aims: To carry out a systematic review between 2018 and 2022 on medicinal plants with potential leishmanicidal activity on parasite strains from the New World causing cutaneous leishmaniasis. Methods: The review study was conducted in four phases following the PRISMA methodology. First, research questions and objectives were formulated to establish the topic areas and construct the search algorithm. Second, a search was performed across different databases, including ScienceDirect, Scopus, PubMed, Web of Science, EBSCO, Taylor and Francis, and Scielo. Third, articles were chosen based on specific inclusion and exclusion criteria. Finally, the relevant information for the review was systematically organized. Results: The search yielded 163 articles, and 12 of them were selected as the basis for the construction of the review. Ethanolic and aqueous extracts stand out, as well as biocompounds such as terpenes and flavonoids. Antioxidant activity on reactive oxygen species was the most frequently cited. Conclusions: Promising terpene and flavonoid molecules with high antileishmanial activity (IC50 &lt;2 μM or &lt;10 μg/mL and SI &gt;1) were identified in this study; these findings provide a scientific basis for the traditional use that communities have given to plants as a therapeutic source to treat cutaneous leishmaniasis in the New World.

Inhibitory effects on bovine babesial infection by iron chelator, 1-(N-acetyl-6-aminohexyl)− 3-hydroxy-2-methylpyridin-4-one (CM1), and antimalarial drugs

BackgroundBabesiosis is an infectious disease caused by protozoa of the apicomplexan phylum, genus Babesia. It is a malaria-like parasitic disease that can be transmitted via tick bites. The apicomplexan phylum of eukaryotic microbial parasites has had detrimental impacts on human and veterinary medicine. There are only a few drugs currently available to treat this disease; however, parasitic strains that are resistant to these commercial drugs are increasing in numbers. Plasmodium and Babesia are closely related as they share similar biological features including mechanisms for host cell invasion and metabolism. Therefore, antimalarial drugs may be useful in the treatment of Babesia infections. In addition to antimalarials, iron chelators also inhibit parasite growth. In this study, we aimed to evaluate the in vitro inhibitory efficacy of iron chelator and different antimalarials in the treatment of Babesia bovis. MethodsCytotoxicity of antimalarial drugs; pyrimethamine, artefenomel, chloroquine, primaquine, dihydroarthemisinine, and the iron chelator, 1-(N-acetyl-6-aminohexyl)− 3-hydroxy-2 methylpyridin-4-one (CM1), were evaluated against Madin Darby Bovine Kidney (MDBK) cells and compared to diminazene aceturate, which is the currently available drug for animal babesiosis using an MTT solution. Afterwards, an evaluation of the in vitro growth-inhibitory effects of antimalarial drug concentrations was performed and monitored using a flow cytometer. Half maximal inhibitory concentrations (IC50) of each antimalarial and iron chelator were determined and compared to the antibabesial drug, diminazine aceturate, by interpolation using a curve-fitting technique. Subsequently, the effect of the drug combination was assessed by constructing an isobologram. Values of the sum of fractional inhibitions at 50% inhibition were then estimated. ResultsResults indicate that all drugs tested could safely inhibit babesia parasite growth, as high as 2500 μM were non-toxic to mammalian cells. Although no drugs inhibited B. bovis more effectively than diminazine aceturate in this experiment, in vitro growth inhibition results with IC50 values of pyrimethamine 6.25 ± 2.59 μM, artefenomel 2.56 ± 0.67 μM, chloroquine 2.14 ± 0.76 μM, primaquine 22.61 ± 6.72 μM, dihydroarthemisinine 4.65 ± 0.22 μM, 1-(N-acetyl-6-aminohexyl)− 3-hydroxy-2 methylpyridin-4-one (CM1) 9.73 ± 1.90 μM, and diminazine aceturate 0.42 ± 0.01 μM, confirm that all drugs could inhibit B. bovis and could be used as alternative treatments for bovine babesial infection. Furthermore, the efficacy of a combination of the iron chelator, CM1, in combination with artefenomel dihydroarthemisinin or chloroquine, and artefenomel in combination with the iron chelator, CM1, dihydroarthemisinin or chloroquine, exhibited synergism against B. bovis in vitro. ConclusionOur evaluation of the inhibitory efficacy of the iron chelator CM1, antimalarial drugs, and a combination of these drugs against B. bovis could be potentially useful in the development and discovery of a novel drug for the treatment of B. bovis in the future.

In vitro and in silico anti-malarial activity and cytotoxicity of n-hexyl 1-O-rutinoside (a glycoside) isolated from Annickia polycarpa (DC.) Setten and Maas ex I.M. Turner (Annonaceae)

Ethnopharmacological relevanceAnnickia polycarpa leaf is an effective anti-malarial agent. However, its chemical constituents have not been isolated and assayed against any pathogen. Aim of the studyTo isolate and characterize anti-malarial compound(s) from the leaf of A. polycarpa. Materials and methodsBioassay-guided fractionation was employed to isolated the compound (AL1) from the chloroform fraction (ALCF) of the basified ethanol extract of A. polycarpa leaf (ALE). AL1 was characterized by LC-MS, 1D and 2D NMR spectroscopic analysis. Anti-malarial activity was evaluated against drug resistance Dd2 and drug sensitive 3D7 Plasmodium falciparum strains using the SYBR green assay. Cytotoxicity and mechanistic studies were determined using tetrazolium-based colorimetric assay and molecular docking respectively. ResultsAL1 was characterized as n-hexyl 1-O-rutinoside. The IC50 values of ALE and ALCF against 3D7 and Dd2 P. falciparum strains ranges from 3.441 (0.3389) - 4.255 (0.2246) μg/mL. The IC50s obtained for n-hexyl 1-O-rutinoside and Artesunate (standard drug) were 7.71 (0.5473) and 0.001 (0.00008) nM against the 3D7 parasite strain respectively. Also, the efficacy of n-hexyl 1-O-rutinoside increased by 24.40% against the chloroquine resistance Dd2 P. falciparum strain whiles that of Artesunate decreased by 98.96%. Furthermore, ALE, ALCF and n-hexyl 1-O-rutinoside were weakly cytotoxic to human RBCs with high selectivity indices. N-hexyl 1-O-rutinoside inhibits P. falciparum chloroquine resistance transporter (PfCRT) and dihydrofolate reductase-thymidylate synthase (PfDHFR-TS) better than chloroquine and pyrimethamine respectively. But, produced similar inhibition of P. falciparum 2-trans-enoyl -ACP-reductase (PfERN) as triclosan. ConclusionThese results show that A. polycarpa leaf and n-hexyl 1-O-rutinoside possessed profound anti-malarial activity and are not cytotoxic. N-hexyl 1-O-rutinoside could therefore, be developed into a new anti-malarial medicine. This is the first study to report the anti-malarial activity of n-hexyl 1-O-rutinoside and its isolation from the genus Annickia.