Search Results (218)

Human cryptosporidiosis caused by the zoonotic apicomplexan parasite Cryptosporidium parvum represents a neglected and re-emerging poverty-related disease. C. parvum possesses minimalistic metabolic capacities and highly depends on its intestinal epithelial host cell for intracellular replication. Based on previous results showing that glycolysis and glutaminolysis inhibition diminished C. parvum replication in vitro, we here investigated the impact of the olive oil component oleocanthal on C. parvum infection in HCT-8 cells under physioxia (5% O₂) and hyperoxia (21% O₂). Oleocanthal targets a broad spectrum of regulatory molecules, amongst which mTOR represents a master regulator of glycolysis and glutaminolysis. Using a host cell pre-treatment as well as a pre- and post-infection treatment protocol, 5 µM oleocanthal reduced C. parvum infection rates between 51% and 94%. Host cellular metabolic conversion rates linked oleocanthal-induced inhibition of C. parvum infection with an impairment in glutaminolysis, representing an important metabolic pathway in intestinal cells. The principal involvement of mTOR in C. parvum inhibition was confirmed by another mTOR-inhibitor (PP242, 0.5 µM), which also reduced C. parvum infection by 70–77%. Given that oleocanthal is not a selective mTOR inhibitor, we assume that this compound drives a multi-target-based inhibition of asexual C. parvum replication, amongst which mTOR is addressed. Full article

Studies spanning decades provide important information about the epidemiology and occurrence of a broad range of diseases that affect the central nervous system (CNS) of dogs. This study analyzed records and formalin-fixed paraffin-embedded (FFPE) CNS tissue samples from necropsied dogs with neuropathologic changes between 1962 and 2022. A total of 134,854 animals, including 20,117 dogs, were submitted for necropsy during this time span. Of these dogs, 2646 displayed alterations of the CNS. Degenerative and non-suppurative inflammatory lesions were the most common changes, accounting for 35.6% and 28.6%, respectively. Vascular diseases, neoplasms, congenital malformations, and suppurative inflammation represented 13.8%, 8.6%, 7.2%, and 5.4% of cases, respectively. Morbillivirus canis, the agent of canine distemper, was the most commonly diagnosed. The second most commonly detected virus, varicellovirus suidalpha1, the agent of pseudorabies, occurred almost exclusively between the mid-1970s and 1990s. Other pathogens, including Lyssavirus rabies, canine herpes virus, tick-borne encephalitis virus, apicomplexan parasites, such as Neospora caninum and Toxoplasma gondii, as well as fungal and other parasitic infections, were less frequently diagnosed. Interestingly, 47.6% of cases with non-suppurative inflammation remained etiologically undetermined. This study provides insights into the epidemiology of canine neurotropic infections and shows the value of FFPE material for investigations of past disease outbreaks. Full article

The conversion from fast-growing tachyzoites to slow-growing bradyzoites is the key factor in establishing the chronic infection and long-term persistence of Toxoplasma gondii. Environmental stressors, such as amino acid starvation and alkaline medium, can trigger the transformation of tachyzoites into bradyzoites. Under such stress conditions, ribosomes slow down, potentially leading to stalling, and ribosomal collisions typically activate ribosome-associated quality control (RQC) pathways. In this study, we investigated the role of T. gondii ribosome quality control complex subunit 2 (TgRqc2), which contains both NFACT and coiled-coil domains, in the parasite’s survival and stage conversion. NFACT represents the “domain” found in the central players involved in RQC, human NEMF and its orthologs FbpA (known as RqcH), Caliban, and Tae2 (known as Rqc2). Phylogenetic analyses revealed that TgRqc2 formed a distinct clade with its orthologs in apicomplexan parasites. The deletion of TgRqc2 impaired T. gondii’s invasion and replication. The Rqc2-knockout strain showed defects in plaque formation and bradyzoite development. Our findings demonstrate that TgRqc2 is essential for T. gondii’s lytic cycle and the conversion of tachyzoites into bradyzoites. RNA-seq analysis further showed that the depletion of TgRqc2 significantly disrupted global transcriptional activity. However, the detailed molecular mechanisms involved remain to be elucidated. In conclusion, our results proved valuable insights that may aid in the development of therapeutic strategies to prevent chronic infection. Full article

Toxoplasma gondii is responsible for the disease toxoplasmosis and has the broadest host range among apicomplexan parasites, as it infects virtually all warm-blooded vertebrates. Toxoplasmosis is a zoonotic and emerging public health concern with considerable morbidity and mortality, especially in the developing world, affecting approximately one-third of the world’s human population. Clinical presentation varies among species, and the infection establishes lifelong chronicity in hosts. Most of the host species (including healthy humans) are asymptomatic on the one hand, it is fatal to marsupials, neotropical primates and some marine mammals on the other hand. In immunocompetent humans, infection is typically asymptomatic, whereas immunocompromised individuals may develop disseminated disease affecting virtually any organ system—most commonly reproductive, cerebral, and ocular systems. Toxoplasmosis spreads by ingestion of food or water contaminated with T. gondii oocysts, consumption of undercooked/raw meat containing tissue cysts, transplacental transmission from mother to fetus, or by receiving infected organ/blood from the infected individual. Toxoplasmosis is mainly diagnosed by serologic tests and polymerase chain reaction (PCR). It is treated with pyrimethamine combined with sulfadiazine or clindamycin, often supplemented with leucovorin, atovaquone, and dexamethasone. Despite having many potent anti-T. gondii antigenic candidates, there is no commercially available vaccine for humans due to many factors, including the complex life cycle of the parasite and its evasion strategies. To date, the only commercially available anti-T. gondii vaccine is for sheep, licensed for veterinary use to prevent ovine abortions. In this review, we have summarized the current understanding of toxoplasmosis. Full article

Babesia divergens is a tick-borne apicomplexan parasite that causes human babesiosis, a malaria-like disease. B. divergens metabolism remains poorly characterized. Here, we employed a multiplatform mass spectrometry-based metabolomics approach (using CE-TOF/MS, GC-QTOF/MS, LC-QTOF/MS, and LC-QqQ/MS) to profile intra- and extracellular metabolic changes in B. divergens-infected and uninfected red blood cells (RBCs) and their supernatants. Our results indicate alterations in the metabolome caused by B. divergens infection and proliferation within RBCs. These findings are consistent with the major metabolic dependencies of B. divergens, including extracellular glucose, glutamine, and arginine, accompanied by the accumulation of glycolytic and TCA cycle intermediates. We identified altered nucleotide metabolism, pentose phosphate pathway activity, and redox imbalance. Depletion of lysoglycerophospholipids, glucose, arginine, and glutamine, and accumulation of free heme and sphingolipids suggested pathogenic effects. Growth experiments indicate that glucose and glutamine, but not hypoxanthine, are required for parasite growth. We additionally discovered a phosphorylated HEPES derivative (PEPES) produced upon B. divergens infection of RBCs in vitro. Collectively, these findings and their global interpretation provide insights into B. divergens metabolism and metabolic dependencies and host–parasite metabolic interactions and outline potential directions for future studies on human babesiosis diagnosis, prognosis assessment, and treatment. Full article

The apicoplast is a highly specialized organelle in the biosynthesis of essential metabolites in most of the apicomplexan protozoa. This organelle is surrounded by four layers of membranes. However, the molecular mechanisms mediating transmembrane transport are not yet fully understood. In this study, we conducted a phenotypic analysis to investigate the role of a major facilitator superfamily transporter (TgApMFS1) in the survival of the parasite. The results indicated that TgApMFS1 is critical for the survival of Toxoplasma gondii in cell culture conditions. Further analysis indicated that these transporters are crucial for the biogenesis of organelles and the metabolic processes of parasite. Full article

Background: Bovine babesiosis, caused by the tick-borne apicomplexan parasite Babesia spp., is an economically significant disease that threatens the cattle industry worldwide. Babesia bovis is the most pathogenic species, leading to high morbidity and mortality in infected animals. One promising approach to vaccination against bovine babesiosis involves the use of multiple protective antigens, offering advantages over traditional live-attenuated vaccines. Tools such as immunobioinformatics and reverse vaccinology have facilitated the identification of novel antigens. Enolase, a “moonlighting” enzyme of the glycolytic pathway with demonstrated vaccine potential in other pathogens, has not yet been studied in B. bovis. Methods: In this study, the enolase gene from two B. bovis isolates was successfully identified and sequenced. The gene, consisting of 1366 base pairs, encodes a predicted protein of 438 amino acids. Its expression in intraerythrocytic parasites was confirmed by RT-PCR. Two peptides containing predicted B-cell epitopes were synthesized and used to immunize rabbits. Hyperimmune sera were then analyzed by ELISA, confocal microscopy, Western blot, and an in vitro neutralization assay. Results: The hyperimmune sera showed high antibody titers, reaching up to 1:256,000. Specific antibodies recognized intraerythrocytic merozoites by confocal microscopy and bound to a ~47 kDa protein in erythrocytic cultures of B. bovis as detected by Western blot. In the neutralization assay, antibodies raised against peptide 1 had no observable effect, whereas those targeting peptide 2 significantly reduced parasitemia by 71.99%. Conclusions: These results suggest that B. bovis enolase contains B-cell epitopes capable of inducing neutralizing antibodies and may play a role in parasite–host interactions. Enolase is therefore a promising candidate for further exploration as a vaccine antigen. Nonetheless, additional experimental studies are needed to fully elucidate its biological function and validate its vaccine potential. Full article

Toxoplasma gondii, a parasitic protozoan, causes zoonotic infections with severe health impacts in humans and warm-blooded animals, underscoring the urgent need for effective vaccines to control these infections. In this study, a DNA vaccine encoding TgROP5, TgROP18, TgGRA7, TgGRA15, and TgMIC6 was formulated using the eukaryotic expression vector pVAX I. IL-24 was delivered as a molecular adjuvant using plasmid pVAX-IL-24. BALB/c, C57BL/6, and Kunming mouse strains received the DNA immunization, after which antibody levels, cytokine production, and lymphocyte surface markers were analyzed to assess immune responses. Additionally, survival rates and brain cyst counts were measured 1 to 2 months post-vaccination in experimental models of toxoplasmosis. As a result, compared to controls, the DNA vaccine cocktail significantly increased serum IgG levels, Th1 cytokine production, and proportions of CD4+/CD8+ T cells, leading to extended survival and reduced brain cyst counts post-challenge with T. gondii ME49. Furthermore, the five-gene DNA vaccine cocktail conferred greater protection compared to single-gene immunizations. Co-administration of IL-24 significantly enhanced the immune efficacy of the multi-gene DNA vaccination. Our findings suggest that IL-24 is an effective molecular adjuvant, enhancing the protective immunity of DNA vaccines against T. gondii, supporting its potential role in vaccine strategies targeting other apicomplexan parasites. Full article

Pre-, pro-, and post-biotics have been used with success in several commercially grown insect species to increase yields and improve health outcomes. However, few studies have been published evaluating the use of nutritional supplements in Acheta domesticus. For this study, we fed day-old, farm-raised, A. domesticus a Saccharomyces cerevisiae postbiotic product at one of three different inclusion rates (0, 0.25, and 0.5%) for 32 days. Crickets were analyzed for differences in average weights, total biomass, percent survival, nutritional analyses, viral qPCR, and 16S/18S microbiomes. Crickets receiving the 0.5% inclusion feed trended towards having a higher total biomass (F = 3.823, p = 0.052) and a higher percent survival per bin (F = 3.667, p = 0.057) than the crickets receiving lower inclusion feeds. No significant differences were found in viral prevalences or loads. Significant changes to the microbiomes were mostly defined by increased abundances of presumed beneficial bacteria (Akkermansia, Catenibacillus, and Odoribacter) in the groups receiving postbiotics rather than by losses of harmful bacteria. For the 18S results, there was an increased abundance of a Gregarian apicomplexan, Leidyana erratica in the treatment groups. Overall, the 0.5% inclusion feed appeared to be beneficial and further study investigating other forms of feed additives is warranted. Full article

Toxoplasma gondii is an Apicomplexan parasite that possesses a well-developed system of scavengers of reactive oxygen species (ROS). Among its components, T. gondii mitochondrial superoxide dismutase (TgSOD2) is essential, as predicted by the CRISPR phenotype index and evidenced by the non-viability of its constitutive knockouts. As an obligate intracellular parasite, TgSOD2 is upregulated during extracellular stages. Herein, we generated a viable TgSOD2 knockdown mutant using an inducible auxin–degron system to explore the biological role of TgSOD2 in T. gondii. Depletion of TgSOD2 led to impaired parasite growth and replication, reduced mitochondrial membrane potential (MMP), abnormalities in the distribution of ATP synthase within its mitochondrial electron transport chain (mETC), and increased susceptibility to mETC inhibitors. Through a proximal biotinylation approach, we identified the interactions of TgSOD2 with complexes IV and V of its mETC, suggesting that these sites are sensitive to ROS. Our study provides the first insights into the role of TgSOD2 in maintaining its mitochondrial redox homeostasis and subsequent parasite replication fitness. Significance: Toxoplasma gondii infects nearly a third of the world population and can cause fetal miscarriages or life-threatening complications in vulnerable patients. Current therapies do not eradicate the parasite from the human hosts, rendering them at risk of recurrence during their lifetimes. T. gondii has a single mitochondrion, which is well-known for its susceptibility to oxidative damage that leads to T. gondii’s death. Therefore, targeting T. gondii mitochondrion remains an attractive therapeutic strategy for drug development. T. gondii’s mitochondrial superoxide dismutase is an antioxidant protein in the parasite mitochondrion and is essential for its survival. Understanding its biological role could reveal mitochondrial vulnerabilities in T. gondii and provide new leads for the development of effective treatments for T. gondii infections. Full article

Rodents are among the most abundant and ecologically diverse mammals, playing key roles in terrestrial ecosystems and often serving as reservoirs for various zoonotic and wildlife pathogens. Among these are protozoan parasites of the genera Hepatozoon and Theileria, which are known to infect a wide range of domestic and wild animals worldwide. However, little is known about the diversity and phylogenetic relationships of these hemoprotozoans in rodent hosts, particularly in the Arabian Peninsula. The aim of this study was to investigate the presence and genetic diversity of Hepatozoon sp. and Theileria sp. in rodents from different regions of Saudi Arabia and to determine potential reservoir species. A total of 111 rodents were captured and identified by molecular analysis of the mitochondrial 16S rRNA gene. Screening for parasites was performed using PCR amplification of the 18S rRNA gene, followed by sequencing, haplotype analysis, and phylogenetic reconstruction using both maximum likelihood and Bayesian inference methods. Our results represent the first molecular detection of Hepatozoon sp. in Arvicanthis niloticus (31.3%), Gerbillus cheesmani (26.5%), G. nanus (28.5%), and Rattus rattus (32.0%) and of Theileria sp. in G. nanus (21.5%) and R. rattus (24.0%) in Saudi Arabia. Haplotype network analysis revealed seven distinct Hepatozoon haplotypes forming a star-like cluster, suggesting host specificity. One divergent haplotype (Hap_2), 19 mutation steps apart, may represent a novel lineage. Phylogenetic analyses grouped Saudi Hepatozoon sequences with those from reptiles and rodents, forming a clade distinct from sequences isolated from felids and canids. In contrast, Theileria sequences showed low diversity, clustering with a single widespread haplotype found in rodents and ruminants in several regions. These findings significantly expand the current knowledge on rodent-associated apicomplexan parasites in Saudi Arabia, revealing novel Hepatozoon haplotypes and highlighting the role of rodents in the transmission of reptile-associated Hepatozoon spp. This study provides basic molecular data crucial to understanding host–parasite relationships and the potential public and veterinary health implications of these parasites in arid ecosystems. Full article

Tick-borne piroplasmids are apicomplexan protozoa that infect a wide range of vertebrate hosts, with significant implications for animal and human health. This study investigated the occurrence and genetic diversity of piroplasmids in wild mammals, unconventional pets, and associated ticks in Goiás state, midwestern Brazil. Between April 2023 and January 2024, 105 blood samples, 22 tissue samples, and 300 ticks were collected from 21 mammalian species housed in wildlife screening centers, zoos, and veterinary clinics. Molecular screening targeting the 18S rRNA gene of piroplasmids detected a 25.7% (27/105) overall positivity, with gray brockets (Subulo gouazoubira) and South American tapirs (Tapirus terrestris) showing the highest infection rates. Three tick samples tested positive, including two Amblyomma sculptum nymphs and a male of Amblyomma dubitatum collected from a tapir and capybara (Hydrochoerus hydrochaeris). Cytauxzoon brasiliensis was reported, for the first time, in cougars (Puma concolor) from Goiás state, midwestern Brazil, indicating the role of this feline as a host of this parasite. Babesia goianiaensis was confirmed in a capybara, and Theileria terrestris in tapirs. Phylogenetic analyses clustered gray brockets-associated Theileria sequences with Theileria sp. previously detected in Neotropical deer from Brazil and Theileria cervi. While the phylogenetic analysis of amino acid sequences of the cytochrome c oxidase subunit III separated Theileria genotypes detected in S. gouazoubira from T. cervi, hsp70-based phylogenetic inferences clustered the genotypes detected in Tapirus terrestris with Theileria terrestris, suggesting host-specific evolutionary lineages. These findings contribute to the understanding of Piroplasmida diversity and circulation in South American wild mammals, emphasizing the need for enhanced molecular surveillance to elucidate transmission dynamics, assess potential health risks, and contribute to the establishment of wildlife conservation and One Health strategies. Full article

The apicomplexan parasite Toxoplasma gondii can potentially infect all warm-blooded animals, including birds, which, due to their high dispersal capabilities, are considered a significant candidate group of sentinel animals that reveal environmental contamination with this protozoan. In the present study, the serologic and molecular prevalences of T. gondii infection were determined in 333 corvids from Romania. Paired meat juice (n = 333) and heart samples (n = 244) were collected and analyzed using the modified agglutination test for antibodies, polymerase chain reaction (PCR) for DNA, and SAG2 molecular marker sequencing for genotyping. The overall T. gondii antibodies prevalence was 19.5%, with 48.1% infected jackdaws, 72.8% rooks, 89.7% hooded crows, 77.5% magpies, and 42.9% jays. Of 244 heart samples analyzed with PCR amplification, only 3 (1.2%) resulted positive and were shown to belong to genotype III through the sequencing of the SAG2 amplicon. This is the first extensive study on T. gondii in crows from Romania. Full article

Piroplasmids (Babesia spp., Rangelia spp., Theileria spp., Cytauxzoon spp.) are tick-borne apicomplexan protozoa that infect, depending on the species, erythrocytes and leucocytes in a wide variety of mammals and birds. The genera Bartonella and Borrelia include vector-borne bacteria that can infect and cause disease in both animals and humans. Detection of hemotropic bacteria and piroplasmids in wild animals is often challenging due to low bacteremia or parasitemia. Digital (d)PCR has proven to be an effective modality for the detection and quantification of DNA of hemotropic pathogens with low parasitemia. This study compared dPCR results from 366 biological samples from seven different Brazilian wild animal groups (5 Xenarthra species, 5 deer species, 3 felid species, 1 canid species, 3 rodent species, 1 bat species, 1 tapir species, and 12 bird species) to two other molecular diagnostic techniques: quantitative real-time (qPCR) and nested (nPCR). For this study, DNA extracted from wild animal blood and spleen samples were subjected to a multiplex dPCR assay for piroplasmids, Bartonella spp., and Borrelia spp. For comparison, the same primers and probes for each agent were used in qPCR assays. Additionally, an nPCR based on the 18S rRNA gene for piroplasmids was performed. The proportions of positive results obtained using dPCR were 85.5% for piroplasmids, 33.6% for Bartonella spp., and 16.7% for Borrelia spp. For all tested agents, dPCR proved to be the technique with the highest sensitivity, making it a useful tool for screening vector-borne agents in biological samples from wild animals with low parasitemia. Full article

Apicortin, a tubulin/microtubule-binding protein, was first described in 2009 as a protein characteristic of apicomplexans; it was found to be present in all Apicomplexa genomes already sequenced. Apart from these, it was found only in Trichoplax adhaerens, the only known representative of Placozoa at the time. Subsequent analyses revealed that it is present in both closely and distantly related taxa of Apicomplexa (Chrompodellids, Squirmids, Dinoflagellates, and Perkinsids, i.e., in Myzozoa). On the other hand, it turned out that it is also present in early-branching fungi that reproduce by zoospores. Now, we have shown that apicortin is found in many deep-branching opisthokonts. In addition to these fungi and T. adhaerens, it is also present in other simple animals, including further Placozoa and Ctenophora, and another opisthokont clade, choanoflagellates. However, apicortin-homologous sequences detected in the genomes/transcriptomes of bilaterian animals are the result of contamination. Full article