Studies on avian haemosporidians in Mexico and around the world reveal poor knowledge about the vectors that transmit avian haemosporidian parasites and their association with vertebrate hosts. Seasonal variations given by changes in temperature and precipitation have significant repercussions on the dynamics of vector-borne infections. The seasonal dynamics of major haemosporidian parasites and their main vectors are mostly unknown. This study aimed to determine the presence of avian haemosporidians in hematophagous mosquitoes and to characterize mosquito-vertebrate host interactions during a year in a peri-urban cloud forest in Xalapa City, Veracruz, Mexico. We analyzed 12 species and a total of 1170 hematophagous mosquitoes. We found the presence of Plasmodium and Haemoproteus genera in the blood meals of mosquitoes. The highest haemosporidian detection and mosquito richness were in June (dry-warm season), whereas the highest mosquito abundance was in October (humid-warm season). We recorded three new haemosporidian lineages in this study. Analysis of blood meals showed that mosquitoes fed on different vertebrate groups (amphibians, reptiles, birds, and humans). This study contributed to the knowledge about the distribution of hematophagous mosquito-haemosporidian-vertebrate host interactions.

Medicinal leeches of the genus Hirudo inhabit large areas of the Palaearctic realm. The distribution range of Hirudo troctina includes the southern Iberian peninsula and the northwestern regions of Africa. H. troctina is used for medical purposes, but only very little is known about the components of its salivary gland secretion. Hirudins, bivalent inhibitors of thrombin, are probably the best known leech-derived bioactive factors. Hirudin-like factors (HLFs) represent another class of salivary gland components that share characteristic genetic and structural markers with hirudins. Hirudin is not a single entity but exists in at least four different variants. However, there are differences among the European members of the genus Hirudo with respect to the actual number of hirudin and HLF genes that are present within their genomes. Here, we describe the identification and molecular cloning of 11 genes that encode for putative hirudin and HLF variants in H. troctina. Three of the genes consist of exons and introns that originate from different "archetype" genes and are likely the result of recombination events. The diversity of hirudin and HLF genes in H. troctina surpasses that of all other European members of the genus Hirudo. The putative hirudin variants and representatives of the HLFs of H. troctina were expressed as recombinant proteins, purified and functionally characterized for their thrombin-inhibiting potencies. Phylogenetic analyses based on hirudin and HLF gene sequences of the leech genera Hirudo, Hirudinaria, and Whitmania supported the hypothesis that hirudins and HLFs diverged early in leech evolution.

Perkinsus marinus, a parasite myzozoa native from the Atlantic Ocean, has spread across various bivalve species in the Pacific Ocean. In the Gulf of California, clams like Larkinia grandis and Leukoma grata have become a secondary fishery, experiencing demand. During annual sampling (August 2017 - July 2018, n = 30/month), the Ray's fluid thioglycollate medium (RFTM) revealed presumptive hypnospores of Perkinsus sp. in L. grandis, with a low parasite prevalence (4.16%) and density (≤ 3.14 hypnospores g tissue). From summer 2020 to summer 2021, P. marinus was seasonally identified in L. grata via staining and PCR tests, showing mean prevalence and density of 48.33% and 48 hypnospores g tissue, respectively. The infection level was light in both clams. This parasite's presence in these new hosts within the southeastern Gulf of California confirms its ability to expand into non-native places, signaling a need for intervention strategies in managing and transporting mollusk species within the region.

Canine visceral leishmaniasis (CVL) is a parasitic disease caused by the protozoan Leishmania infantum. Neurological infection occurs due to the parasite's ability to cross the blood-brain barrier. It is known that dogs can remain infected with a subclinical infection for life, potentially acting as reservoirs for L. infantum when bitten by sandflies. In this context, the objective of this study was to evaluate the occurrence of Leishmania spp. in the brains of dogs from the metropolitan region of Rio Grande do Sul, Brazil, without a history of neurological disease but residing in an endemic area for L. infantum. A total of 200 samples, from 2022 to 2023, were evaluated using conventional Polymerase Chain Reaction (PCR) with the primers Leishmini-F GGKAGGGGCGTTCTGC and Leishmini-R STATWTTACACCAACCCC, aiming to amplify a product of 120 base pairs for Leishmania spp. To identify the species, a multiplex PCR was used, differentiating L. braziliensis (127 bp), L. amazonensis (100 bp), and L. infantum (60 bp), with the molecular target being the conserved region of the kinetoplast DNA (kDNA) minicircle, specific to Leishmania spp. Of the 200 samples evaluated, 26.5% (53/200) tested positive in the conventional PCR reaction for Leishmania spp., with the PCR multiplex the only species detected was Leishmania infantum. The average age of the positive animals was 5.08 years, with 47.2% being females and 52.8% being males; among these, mixed-breed dogs were the most predominant, representing 43.4% of the total. Clinical signs varied: hepatomegaly in two dogs, pronounced neutrophilic hepatitis in one, splenomegaly in one with lymphoid hyperplasia, and glomerulonephritis and nephritis in two animals. Mild anemia and thrombocytopenia were found in eight, with pale mucous membranes in three, and diffuse alveolar edema in one case. Notable pathological findings included suspected distemper in one animal and lymphoplasmacytic meningitis in another. Histopathological findings revealed alveolar edema and acute renal failure. A third dog exhibited bilateral hydrocephalus and diffuse edema in the brain. Additional changes, such as mild inflammatory infiltrate and slight vacuolar degeneration, were observed in 11.3% of the analyzed brains. There was no clinical suspicion of leishmaniasis in any of the studied cases. Therefore, the detection of L. infantum DNA in the brains of dogs suggests that animals with subclinical infection may play a crucial role in the spread of leishmaniasis, and infection by Leishmania spp. should be considered as a differential diagnosis for neurological disease in endemic areas for the protozoan.

NLRP3 inflammasome-mediated inflammatory responses play pivotal functions in innate immunity. However, its homeostatic regulation still needs to be better understood. Here we explore the effect and potential mechanism of IL-33 on NLRP3 inflammasome upon Toxoplasma gondii infection through a series of molecular biology and immunological experiments, including western blot, qRT-PCR, and ELISA. We demonstrated that T. gondii infection induces the expression of IL-33, and IL-33-deficient (IL-33) mice exhibit longer survival time than wild-type (WT) mice upon T. gondii infection. IL-33 deficiency promotes the expression of NLRP3 and ASC and the secretion of IL-1β, while exogenous IL-33 inhibits NLRP3 inflammasome. Furthermore, T. gondii infection results in the M2 polarization of macrophages, exacerbated by exogenous IL-33, which also promotes the proliferation of T. gondii. These findings showed that IL-33 deficiency attenuates T. gondii infection by promoting NLRP3 inflammasome, advancing the understanding of the role of IL-33 in inflammation.

Acanthamoeba castellanii is the causative pathogen of a severe eye infection, known as Acanthamoeba keratitis and a life-threatening brain infection, named granulomatous amoebic encephalitis. Current treatments are problematic and costly and exhibit limited efficacy against Acanthamoeba parasite, especially the cyst stage. In parallel to drug discovery and drug repurposing efforts, drug modification is also an important approach to tackle infections, especially against neglected parasites such as free-living amoebae: Acanthamoeba. In this study, we determined whether modifying pentamidine and doxycycline through chitosan-functionalized graphene oxide loading enhances their anti-amoebic effects. Various concentrations of doxycycline, pentamidine, graphene oxide, chitosan-functionalized graphene oxide, and chitosan-functionalized graphene oxide loaded with doxycycline and pentamidine were investigated for amoebicidal effects against pathogenic A. castellanii belonging to the T4 genotype. Lactate dehydrogenase assays were performed to determine toxic effects of these various drugs and nanoconjugates against human cells. The findings revealed that chitosan-functionalized graphene oxide loaded with doxycycline demonstrated potent amoebicidal effects. Nanomaterials significantly (p < 0.05) inhibited excystation and encystation of A. castellanii without exhibiting toxic effects against human cells in a concentration-dependent manner, as compared with other formulations. These results indicate that drug modifications coupled with nanotechnology may be a viable avenue in the rationale development of effective therapies against Acanthamoeba infections.

Egress represents a crucial process employed by Neospora caninum in the establishment of infection. Dense granule proteins (GRAs), secreted by the dense granule, play significant roles in modifying the parasitophorous vacuole, maintenance of morphology, and regulating host-cell interactions. However, their precise involvement in tachyzoite egress remains inadequately characterized. In this study, we identified a homologous gene, Ncgra41, corresponding to the dense granule protein 41 (GRA41) of Toxoplasma gondii, which is associated with egress, utilizing NCBI and ToxoDB databases. NcGRA41 is localized extracellularly within dense granules and intracellularly within parasitic vacuoles. Deletion of NcGRA41 did not affect tachyzoites invasion or proliferation but significantly reduced egress capacity and pathogenicity in mice. The phenotypic characteristics were restored in a complementary strain. Further investigation revealed that the absence of NcGRA41 reduced gliding motility and the transcription level of the subtilisin-like protein (SUB1). A microneme secretion assay demonstrated a significant decrease in NcMIC1 secretion, along with reduced expression levels of NcMIC1, NcMIC4, and NcMIC8. These findings demonstrate that NcGRA41, a novel dense granule protein in N. caninum, modulates tachyzoites egress and influences pathogenicity by regulating microneme secretion.

In Xinjiang, previous reports of cystic echinococcosis (CE) in livestock have focused on West and Middle Tianshan Mountains. In contrast, there is an absence of research on CE in the East Tianshan Mountains. To determine the epidemiology and genetic diversity of Echinococcus granulosus sensu stricto (s.s.) in domestic animals in the East Tianshan Mountains, Xinjiang, China, the livers and lungs of 1773 domestic animals were examined, between January 2023 and March 2023, and 40 cysts were collected. Polymerase chain reaction was used for the molecular diagnosis of the cysts. Statistical results showed that the overall prevalence of echinococcosis in sheep was 6.92% (114/1646), which was significantly higher than that in cattle (1.57%, 2/127). A total of 40 cyst isolates were obtained, including 38 from sheep and 2 from cattle. Genomic DNA was extracted, and the mitochondrial cytochrome c oxidase subunit 1 (cox1) gene was amplified by PCR to obtain the target 850 bp fragment. The results revealed that all the isolates had the G1 genotype, with similarities ranging from 98.88-100%. Haplotype network analysis revealed 32 haplotypes of the cox1 gene, among which Hap_7 was the main haplotype. Furthermore, haplotype diversity (Hd = 0.313 ± 0.093) and nucleotide diversity (π = 0.00173 ± 0.00079) were lower in the East Tianshan Mountains than in other regions, indicating that the populations are genetically less differentiated. Tajima's D and Fu's Fs tests were negative (p ≤ 0.01), indicating an expansion of the population in the East Tianshan Mountains of Xinjiang, similar to the results previously reported for Xinjiang. The low fixation index (Fst) ranged from negative values (Gansu Province) to 0.30346 (Mongolia), indicating that the genetic differentiation between the East Tianshan Mountains and Gansu, and Tibet, Xinjiang was relatively low, with frequent gene flow. In this survey, two 'new' haplotypes were identified in the East Tianshan livestock. In addition, two different haplotypes of liver and lung infections were found in one cattle. This survey provides information on the epidemiology and genetic diversity of E. granulosus s.s. in the East Tianshan Mountains of Xinjiang, China.

Invertebrate mitogenomes are generally fixed with formal 37 genes: 13 PCGs encoded subunits of OXPHOS, 2 ribosomal RNA (rRNA) functional in the translation of these PCGs and 22 transfer RNA (tRNA) genes. The order of these genes varies greatly among organisms and named rearrangement. Rearrangement patterns of mitochondrial genomes may shed light on mutation processes and evolutionary relationships of organisms. Mitochondrial gene organization is highly variable among Acari, so rearrangement is a very common mitogenomic pattern in this group. In this study, 258 unique Acari (Acariformes + Parasitiformes) mitogenomes were downloaded from NCBI and studied about rearrangement patterns. Sixty-seven mitotypes were determined among Acari and the most rearranged genes were trnL1 and nad2. Following that, trnI, trnS1, trnN, trnE, trnT, and trnP genes are remarkably mobile (RF > 95%). Conversely, atp6, cox3, trnG, and cytb genes also appears to be quite stable (RF < 20%). Within Acari, mean distance calculations are varied from 1.210 in atp8 to 0.155 in rrnS. Contrary to expectations, among Acari mobile tRNA genes appear to be conserved in sequences, whereas PCGs have higher distance values and seem to be mutated. Consistently, tRNA genes seem saturated, but some PCGs (atp6, cox genes, cytb, nad1, and nad6) are not saturated. These values do not correlate with each other (p > 0.005). This discrepancy may indicate that the genes were rearranged after mutation load; consistent with this, DAMBE saturation values are also not correlated with RF values. Parasitiformes mitogenomes are more mobile than Acariformes mitogenomes and may be under the effect of selective sweeping.

Haematophagous leeches rely on a broad variety of bioactive compounds to secure a sufficient blood meal from their vertebrate prey. Both the primary (platelet aggregation) and secondary (blood coagulation) haemostasis are the main targets of action. The platelet aggregation inhibitor decorsin was first described in the North American leech, Macrobdella decora Say, 1824, whereas the bivalent thrombin inhibitor hirudin was originally identified in the European medicinal leech, Hirudo medicinalis Linnaeus, 1758. Hirudin blocks both the catalytic site and the fibrinogen-binding site (exosite I) of thrombin. Haemadin of the Indian land leech Haemadipsa sylvestris Blanchard, 1894, is also a highly efficient bivalent thrombin inhibitor but blocks exosite II of thrombin. So far, only the archetypal form of haemadin from H. sylvestris has been purified and functionally characterized, and two putative haemadins have been identified in the salivary transcriptome of Haemadipsa interrupta Moore, 1835, a terrestrial leech inhabiting mainly the Malayan peninsula. Here, we describe the identification of ten additional putative haemadins in the transcriptomic data set of H. interrupta, first generated by another study. Furthermore, we identified a putative oligomeric decorsin, which represents the first finding of this anticoagulant in a haemadipsid leech. Both the putative decorsin and a selection of haemadins were expressed, purified, and functionally characterized. The putative haemadins displayed a broad spectrum of thrombin-inhibitory potencies, whereas the putative oligomeric decorsin was indeed a weak inhibitor of platelet aggregation.

Babesia spp. and Theileria spp. are tick-borne apicomplexan protozoa that can cause disease in animals and humans. Deer are considered reservoirs for a wide variety of Piroplasmida species, including some potentially zoonotic. This study aimed to investigate the occurrence and genetic diversity of piroplasmids in wild deer sampled in four Brazilian states (São Paulo, Mato Grosso do Sul, Paraná and Goiás). For this purpose, extracted DNA samples from 181 deer buffy coat samples (138 Blastocerus dichotomus, 26 Subulo gouazoubira, 4 Mazama jucunda, 3 Mazama rufa and 10 Ozotocerus bezoarticus) were subjected to a nested PCR (nPCR) assay based on the 18S rRNA gene in order to perform a screening for piroplasmids and characterized based on the near-complete 18S rRNA, hsp70 and cox-3 genes. As a result, 75.14% (136/181) samples were positive for piroplasmids. Of these, 108 (79.41%), 101 (74.26%) and 67 (49.26%) were positive to near complete 18S rRNA, hsp70 and cox-3 genes, respectively. Phylogenetic analyses based on three molecular markers showed similar topology to each other. All sequences obtained in the present study were positioned into the Theileria sensu stricto clade, forming a distinct clade, albeit close to T. cervi. Most sequences grouped together into a large clade divided into subclades, which were often related to deer genus/species, showing that Theileria lineages seemed to show specificity according to deer genus/species. Two 18S rRNA sequences (one obtained from S. gouazoubira and another from M. jucunda) were positioned into a different clade, apart from other sequences detected in this study, indicating that different species of Theileria occur in deer from Brazil. Two subclusters were observed in the phylogenetic analysis based on the hsp70 gene: the first containing only sequences detected in marsh deer and the second grouping sequences detected in brocket deer (Mazama spp. and S. gouazoubira). The latter was also divided into smaller clades that grouped Theileria genotypes according to deer species (M. jucunda, M. rufa and S. gouazoubira). This study provides the first molecular evidence of Theileria infection in M. jucunda, as well as co-infection by distinct Theileria (sub)species/genotypes in the same deer was evidenced. Finally, this study expanded the knowledge on the diversity of Theileria spp. infecting deer from South America.

Toxoplasma gondii and Neospora caninum infect a wide range of domestic and wild animals, including humans, in the case of T. gondii, and cause economic losses in livestock due to abortion and neonatal mortality. In Somalia, zoonotic diseases are concerning due to cultural practices and livestock's economic importance, but surveillance is limited. This study aimed to determine the seroprevalence and molecular prevalence of T. gondii and N. caninum in Somali sheep, goats, and cattle. A cross-sectional study was conducted between December 2018 and January 2020 in Benadir and Lower Shabelle regions of Somalia. Blood samples were collected from 128 cattle, 184 goats, and 46 sheep. Serum samples were tested for anti-T. gondii and anti-N. caninum antibodies using IFAT, and PCR was performed on extracted DNA to detect T. gondii and N. caninum DNA. Overall, 106/358 (29.6%) animals tested positive for anti-T. gondii antibodies, with the highest prevalence in sheep (62.5%), followed by goats (30.4%) and cattle (15.6%) (P < 0.001). For anti-N. caninum antibodies, 13/358 (3.6%) animals tested positive, with cattle showing the highest prevalence (6.2%), followed by goats and sheep (both 2.2%). Co-seropositivity for both antibodies was found in cattle and sheep. Molecular detection of T. gondii DNA revealed a prevalence of 9/358 (2.5%), primarily in sheep (15.2%) and cattle at 1.6% while all goat samples tested negative. No samples were positive for the N. caninum Nc5 gene. This study reveals T. gondii and N. caninum prevalence in Somali ruminants, highlighting the need for better surveillance and control.

Asymptomatic carriers of Plasmodium falciparum represent important parasite reservoirs maintaining malaria transmission in the community. This study aimed on the one hand to screen the other household members living with children under seasonal malaria chemoprevention (SMC) coverage in order to determine the level of malaria infection in this population and on the other hand to determine the appropriate type of rapid diagnostic test (RDT) for this screening to detect these asymptomatic carriers in the community. During the 2022 SMC campaign (July to October), a cross-sectional survey was carried out in 745 participants who were screened by ultrasensitive rapid diagnostic test (usRDT), standard rapid diagnostic test (rRDT) and microscopy. Out of them, 395 had microscopy results available and were included in the data analysis. The prevalence of asymptomatic carriers of asexual forms of Plasmodium falciparum was 26.58% (105/395) while sexual forms were found in 5.32% (21/395) of the study population. Children from 5 to 15 years had the highest prevalence of P. falciparum asexual forms 35.76% (59/165) compared with older participants. Malaria positivity rate for rRDT and usRDT was 29.40% (219/745) and 40.49% (305/745) respectively. The usRDT had a higher sensitivity than the rRDT (72.38% (95% CI 62.8-80.66) vs. 60.95% (95% CI 50.94-70.33)). In terms of specificity, rRDT had a higher specificity 82.41% (95% CI 77.53-86.62) versus 69.66% (95% CI 64.01-74.89) for usRDT. This study reports a high prevalence of parasite carriers in household members of children under SMC coverage in Nanoro, Burkina Faso. In conclusion, usRDT seems more appropriate for strategies based on detection and treatment of parasite carriers within the community.

Fish are exposed to various parasites such as monogeneans which are flatworms that preferentially affect the gills and skin of these hosts. This study represents the first investigation concerning the distribution of monogenean infracommunities of Enteromius guirali in Cameroon. A total of 100 fish were collected from Cesala River and preserved in 8% formalin solution. Parasite community consisted of Dactylogyrus mendehei and D. nyongensis. These monogenean species adopted an aggregated distribution. Their mean intensities and prevalence increased with host size. Both sides of E. guirali were similarly infested, and no preference for gill arches was recorded for each parasite species. Posterior hemibranch was always the most parasitized, showing the highest parasitic load. Median sector and distal filamentous zone were highly colonized by both monogenean species. This study contributes to understand that the large gill surface offered by great hosts, heterogeneity, bilateral symmetrically of the host's gill system on one hand, and the morphology of haptoral parts and reproduction of parasites, on the other hand, are the main factors which determine the distribution of studied component community.

Ovine toxoplasmosis caused by Toxoplasma gondii is a zoonotic disease responsible for triggering reproductive problems in sheep with great public health impact. The zoonotic risk for humans occurs through the ingestion of meat contaminated with bradyzoites, water and vegetables contaminated with oocysts, or by ingesting unprocessed sheep milk contaminated with tachyzoites. The study aimed to determine the seroprevalence, isolate, and genetically characterize T. gondii from sheep in northeastern Brazil. Blood and tissue samples, including the liver, heart, brain, lung, and diaphragm, were collected from 256 sheep in slaughterhouses. The presence of anti-T. gondii IgG antibodies was assessed by the indirect fluorescent antibody test (IFAT), and sera with titers ≥ 1: 64 were considered positive. Tissue samples from seropositive sheep were digested in acidic pepsin solution and inoculated into two Swiss mice for T. gondii isolation and further characterized by molecular methods. The prevalence of anti-T. gondii antibodies in sheep was 31.64% and the isolation rate was 40.7%. Sixteen out of 33 isolates were submitted to mnRFLP-PCR using ten genetic markers. Four isolates were fully genotyped, and four isolates were amplified at least in one marker. All isolates were characterized as atypical genotypes, and ToxoDB #57 (n = 3) and #143 (n = 1) were identified. The detection of viable T. gondii in sheep from slaughterhouses in Brazil highlights the risk of human exposure. There is the need for proactive control and prevention measures to mitigate the risk of human exposure, considering the importance of sheep meat as a source of protein to humans.

A case of polyparasitism in an 8-year-old female cat from Southern Brazil is reported. Among the described clinical signs, both gurltiosis and lagochilascariasis are of rare occurrence, being categorized as neglected diseases. The diagnosis of feline crural parasitic paraplegia was established through clinical signs, necropsy lesion observations, and the presence of Gurltia paralysans in histological sections of the spinal cord. Lagochilascaris minor was diagnosed with an ulcerated wound in the cervical region and further confirmed by coprology. In addition, other gastrointestinal parasites (i.e., Trichuris spp., Diphyllobothriidae, Toxocara cati, Ancylostoma spp., Cystoisospora spp., and Entamoeba spp.) were diagnosed through fecal examination and Sarcocystis felis by molecular analysis of muscle cysts. Aelurostrongylus abstrusus was detected in histological sections of lung parenchyma and confirmed by molecular analyses. The multiple parasitism detected in this case represents a unique report in terms of the number of parasites in the same individual, indicating the risk represented by stray cats as spreaders of zoonotic agents.