First report of Echinococcus granulosus genotype 1 in a wild boar (Sus scrofa) from China.

Echinococcosis is a worldwide disease endemic to the western region of China. In 2023, echinococcosis was detected in one of 27 wild boars (Sus scrofa) in Yili Prefecture, Xinjiang, northwestern China. Histopathological staining and full sequence mitochondrial (mt) analysis were used to determine the infection genotype. Echinococcus granulosus was detected in the wild boar liver, and the cystic lesion characteristics indicated the E. granulosus genotype (G1). This case is the first confirmation of wild boar serving as a transmitter for the G1 genotype of E. granulosus within China. These findings suggest that surveillance is needed to assess the risk of E. granulosus sensu lato transmission to humans and wild animals.

Molecular phylodiagnosis of Echinococcus multilocularis and Echinococcus granulosus among canids in Guilan province, northern Iran.

Canine-transmitted worms and an uncontrolled deworming program of stray dogs have caused that accurate diagnosis of zoonotic parasites received notable attention in endemic regions. This study aimed to assess the presence of E. multilocularis and E. granulosus infections in canids from Guilan province, northern Iran. A total of 272 fecal samples from carnivores were collected across 24 different regions in Guilan province between 2023 and 2024. All fecal specimens were observed following concentration and flotation techniques. DNAs of taeniid eggs were extracted, amplified, and sequenced targeting of specific mitochondrial Cox1 gene for E. granulosus and NAD 1 gene for E. multilocularis. On the base of molecular and phylogenetic analysis 1.47 % (in jackal) and 25 % (in dogs and jackal) of samples were positive for E. multilocularis and E. granulosus sensu strico G1 genotype, respectively. Molecular technique was found to be more sensitive in detecting infection in comparison with conventional techniques. Sequence analysis of Cox1 indicated a high genetic diversity (Haplotype diversity; 0.933; Number of haplotypes, h: 7) in E. granulosus G1. Current findings show that canids particularly jackals play potential role of definitive host in maintenance and transmission dynamic of E. multilocularis and E. granulosus in northern Iran. The presence of these infections is of particular concern in Guilan province due to the high influx of tourists, increasing the risk of transmission to humans. Therefore, the implementation of preventive programs is warranted to apply hygienic practices and adjusting deworming programs for the canids and at-risk individuals in the region.

Immunization with a Mu-class glutathione transferase from Echinococcus granulosus induces efficient antibody responses and confers long-term protection against secondary cystic echinococcosis.

Cystic echinococcosis, a zoonosis caused by cestodes belonging to the Echinococcus granulosus sensu lato (s.l.) genetic complex, affects humans and diverse livestock species. Although a veterinary vaccine exhibiting high levels of antibody-mediated protection has successfully reached the market, the large genetic diversity among parasite isolates and their particular host preferences, makes still necessary the search for novel vaccine candidates. Glutathione transferases (GSTs) constitute attractive targets for immunoprophylaxis due to their outstanding relevance in helminth detoxification processes, against both exogenous and endogenous stressors. Among the six GSTs known to be expressed in E. granulosus s.l., EgGST1 (Mu-class), EgGST2 (Sigma-class), and EgGST3 (a still non-classifiable isoenzyme), show the highest proteomic expression. Therefore, their recombinant forms -rEgGST1, rEgGST2 and rEgGST3- were herein analyzed regarding their potential to induce long-term antiparasite protection in mice. Only immunization with rEgGST1 induced long-lasting protection; and accordingly, rEgGST1-specific antibodies enhanced the parasite killing through both the classical activation of the host complement system and the antibody-dependent cellular cytotoxicity by macrophages. These results support further testing of rEgGST1 as a vaccine candidate in diverse hosts due to the broad expression of EgGST1 in different parasite stages and tissues.

The monodomain Kunitz protein EgKU-7 from the dog tapeworm Echinococcus granulosus is a high-affinity trypsin inhibitor with two interaction sites.

Typical Kunitz proteins (I2 family of the MEROPS database, Kunitz-A family) are metazoan competitive inhibitors of serine peptidases that form tight complexes of 1:1 stoichiometry, mimicking substrates. The cestode Echinococcus granulosus, the dog tapeworm causing cystic echinococcosis in humans and livestock, encodes an expanded family of monodomain Kunitz proteins, some of which are secreted to the dog host interface. The Kunitz protein EgKU-7 contains, in addition to the Kunitz domain with the anti-peptidase loop comprising a critical arginine, a C-terminal extension of ∼20 amino acids. Kinetic, electrophoretic, and mass spectrometry studies using EgKU-7, a C-terminally truncated variant, and a mutant in which the critical arginine was substituted by alanine, show that EgKU-7 is a tight inhibitor of bovine and canine trypsins with the unusual property of possessing two instead of one site of interaction with the peptidases. One site resides in the anti-peptidase loop and is partially hydrolyzed by bovine but not canine trypsins, suggesting specificity for the target enzymes. The other site is located in the C-terminal extension. This extension can be hydrolyzed in a particular arginine by cationic bovine and canine trypsins but not by anionic canine trypsin. This is the first time to our knowledge that a monodomain Kunitz-A protein is reported to have two interaction sites with its target. Considering that putative orthologs of EgKU-7 are present in other cestodes, our finding unveils a novel piece in the repertoire of peptidase-inhibitor interactions and adds new notes to the evolutionary host-parasite concerto.

Echinococcus granulosus comparative genotyping in sheep in Saudi Arabia and Egypt.

Background: Cystic echinococcosis (CE), which is triggered by the parasite Echinococcus granulosus, is a global zoonotic disease that is common in rural regions in which there are frequent encounters between dogs and other domestic animals. The disease can have devastating consequences, impacting the health of people and animals and leading to huge financial losses, especially in the agricultural industry. In the Kingdom of Saudi Arabia (KSA) and Egypt, despite the high incidence of disease, few investigations have been conducted into the genetic variation in species of the genus Echinococcus. Aim: This study sought to compare the genetic features of the hydatid cysts carried in sheep in KSA with those found in Egypt. Methods: DNA from the protoscolices was used in a PCR targeting the mitochondrial NADH dehydrogenase 1 (NAD1), cytochrome c oxidase subunit 1 (COX1), and nuclear actin II (ACT II) genes, and the resulting amplification products of 30 KSA and Egyptian isolates were sequenced and compared. Results: Among the sheep in KSA, the overall prevalence of CE was 0.51%. Of the sheep cyst DNA samples, 95%, 100%, and 52% were positive for the Cox1, nad1, and act II genes, respectively. Targeting all three genes, all KSA samples belonged to the E. granulosus genotype (G1), whereas all Egyptian isolates belonged to E. granulosus (G1) and E. canadensis (G6). Conclusion: We conclude that isolates of E. granulosus from the two countries shared a common origin in Arabic North Africa, with sheep and camels as common hosts.

Echinococcus granulosus Sensu Stricto and Echinococcus multilocularis in a Gray Wolf (Canis lupus) in Turkey: Further Evidence for Increased Risk of Alveolar Echinococcosis in Urban Areas.

OBJECTIVE: The aim of this study was to identify Echinococcus species by morphological and molecular means. METHODS: A dead gray wolf (Canis lupus) was found near Erzurum province and brought to the parasitology laboratory. Sedimentation and counting technique (SCT) and polymerase chain reaction (PCR) analysis were conducted. RESULTS: The SCT implications indicated that the wolf had a substantial worm burden (62,720 and 49,280 parasites) due to a co-infection of E. granulosus s.l. and E. multilocularis. Genus/species-specific PCR was used to analyze DNA extracted from adult worms and confirmed as E. granulosus s.s. and E. multilocularis, utilizing COI and 12S rRNA gene sequence analysis, respectively. CONCLUSION: This report presents the first co-detection of E. granulosus s.s. and E. multilocularis in a gray wolf found in an urban area in a highly endemic area for human echinococcosis in northeastern Turkey. The results emphasize that AE is not only a problem of rural areas, but also occurs in urban areas, which may pose a threat to public health. Therefore, surveillance in urban areas is crucial. The need to develop new control strategies for domestic and wildlife in the study area is also highlighted.

Echinococcus granulosus ubiquitin-conjugating enzymes (E2D2 and E2N) promote the formation of liver fibrosis in TGFß1-induced LX-2 cells.

BACKGROUND: Cystic echinococcosis (CE) is a widespread zoonosis caused by the infection with Echinococcus granulosus sensu lato (E. granulosus s.l.). CE cysts mainly develop in the liver of intermediate hosts, characterized by the fibrotic tissue that separates host organ from parasite. However, precise mechanism underlying the formation of fibrotic tissue in CE remains unclear. METHODS: To investigate the potential impact of ubiquitin-conjugating enzymes on liver fibrosis formation in CE, two members of ubiquitin-conjugating (UBC) enzyme of Echinococcus granulosus (EgE2D2 and EgE2N) were recombinantly expressed in Escherichia coli and analyzed for bioinformatics, immunogenicity, localization, and enzyme activity. In addition, the secretory pathway and their effects on the formation of liver fibrosis were also explored. RESULTS: Both rEgE2D2 and rEgE2N possess intact UBC domains and active sites, exhibiting classical ubiquitin binding activity and strong immunoreactivity. Additionally, EgE2D2 and EgE2N were widely distributed in protoscoleces and germinal layer, with differences observed in their distribution in 25-day strobilated worms. Further, these two enzymes were secreted to the hydatid fluid and CE-infected sheep liver tissues via a non-classical secretory pathway. Notably, TGFß1-induced LX-2 cells exposed to rEgE2D2 and rEgE2N resulted in increasing expression of fibrosis-related genes, enhancing cell proliferation, and facilitating cell migration. CONCLUSIONS: Our findings suggest that EgE2D2 and EgE2N could secrete into the liver and may interact with hepatic stellate cells, thereby promoting the formation of liver fibrosis.

Modulatory actions of Echinococcus granulosus antigen B on macrophage inflammatory activation.

Cestodes use own lipid-binding proteins to capture and transport hydrophobic ligands, including lipids that they cannot synthesise as fatty acids and cholesterol. In E. granulosus s.l., one of these lipoproteins is antigen B (EgAgB), codified by a multigenic and polymorphic family that gives rise to five gene products (EgAgB8/1-5 subunits) assembled as a 230 kDa macromolecule. EgAgB has a diagnostic value for cystic echinococcosis, but its putative role in the immunobiology of this infection is still poorly understood. Accumulating research suggests that EgAgB has immunomodulatory properties, but previous studies employed denatured antigen preparations that might exert different effects than the native form, thereby limiting data interpretation. This work analysed the modulatory actions on macrophages of native EgAgB (nEgAgB) and the recombinant form of EgAg8/1, which is the most abundant subunit in the larva and was expressed in insect S2 cells (rEgAgB8/1). Both EgAgB preparations were purified to homogeneity by immunoaffinity chromatography using a novel nanobody anti-EgAgB8/1. nEgAgB and rEgAgB8/1 exhibited differences in size and lipid composition. The rEgAgB8/1 generates mildly larger lipoproteins with a less diverse lipid composition than nEgAgB. Assays using human and murine macrophages showed that both nEgAgB and rEgAgB8/1 interfered with in vitro LPS-driven macrophage activation, decreasing cytokine (IL-1ß, IL-6, IL-12p40, IFN-ß) secretion and ·NO generation. Furthermore, nEgAgB and rEgAgB8/1 modulated in vivo LPS-induced cytokine production (IL-6, IL-10) and activation of large (measured as MHC-II level) and small (measured as CD86 and CD40 levels) macrophages in the peritoneum, although rEgAgB8/1 effects were less robust. Overall, this work reinforced the notion that EgAgB is an immunomodulatory component of E. granulosus s.l. Although nEgAgB lipid's effects cannot be ruled out, our data suggest that the EgAgB8/1 subunit contributes to EgAgB´s ability to regulate the inflammatory activation of macrophages.

Effects of annexin B18 from Echinococcus granulosus sensu lato on mouse macrophages.

Cystic echinococcosis (CE) is a zoonotic disease, caused by Echinococcus granulosus sensu lato (E. granulosus s. l.), which posed significant public health concern globally. E. granulosus s. l. annexin B18 (EgANXB18) acts as a secretory protein, exerting a crucial influence in mediating host-parasite interactions. Recombinant annexin B18 (rEgANXB18) was expressed by Escherichia coli and the immunoreactivity was assessed by western blotting. The binding affinity between rEgANXB18 and total protein of RAW264.7 cells was assessed by ELISA. The impact of rEgANXB18 on the metabolic activity of RAW264.7 cells was assayed by Cell Counting Kit-8 assay. The mRNA levels of polarization markers (inducible nitrous oxide synthase (iNOS) and arginase 1 (Arg1)) and key cellular factors (IL-1ß，IL-6，IL-10 and TNFα) were evaluated by qRT-PCR. rEgANXB18 was successfully expressed and recognized by E. granulosus s.l. infected canine sera, as well as could bind to the total protein of RAW264.7 cells. Additionally, rEgANXB18 could promote metabolic activity at 5, 10, 20, and 40 µg/mL while no significant impact on metabolic activity was observed at 80 µg/mL. Co-culture RAW264.7 cells with rEgANXB18 resulted in significantly upregulation of the transcript levels of polarization markers iNOS and Arg1. Moreover, rEgANXB18 significantly upregulated the transcript levels of IL-1ß, IL-6, TNFα, and IL-10, while dose-effect relationship was observed in IL-1ß, IL-6, and IL-10. Our results indicated that EgANXB18 showed the potential to regulate immune response of macrophages by shifting the cell polarization and cytokine profile, thereby promoting the parasitism of CE.

Cystic echinococcosis in cattle and sheep caused by Echinococcus granulosus sensu stricto genotypes G1 and G3 in the USA.

BACKGROUND: Endemic domestic dog-ruminant cycles and human cystic echinococcosis caused by Echinococcus granulosus have been sporadically reported in the United States. However, there is a paucity of molecular data describing the genotypes and haplotypes of this important cestode in domestic ruminant hosts. METHODS: Ninety-four cysts from the lungs and/or livers of slaughtered beef cattle (76 samples), dairy cows (five samples) and sheep (13 samples) were collected from abattoirs in four states of the USA. Samples were genotyped at two mitochondrial loci, cox1 and nad5. Sequences were used to determine species, genotypes and haplotypes using median joining networks and Bayesian phylogenetic analyses. Cyst fertility was assessed in hematoxylin and eosin-stained sections. Additionally, previously reported autochthonous E. granulosus infections in the USA in various hosts were mapped. RESULTS: Based on cox1 sequences obtained from 94 cysts, 89 (94.7%) were identified as E. granulosus G1/G3, while five (5.3%) were Taenia hydatigena. Taenia hydatigena were only isolated from sheep. Based on nad5 sequences obtained from 89 hydatid cysts, 96.6% and 3.4% belonged to E. granulosus sensu stricto genotypes G1 and G3 respectively. Two haplotypes were found among E. granulosus cox1 sequences, neither of which was geographically unique. Six haplotypes were found among nad5 sequences in genotype G1, of which five were novel, while one haplotype was found in genotype G3. In the concatenated cox1-nad5 dataset, seven haplotypes were identified, of which six were geographically unique. All cysts from cattle were non-fertile. Four cysts from sheep were fertile. CONCLUSIONS: All genotyped samples belonged to E. granulosus s.s. This is the first study to our knowledge to confirm the presence of genotypes G1 and G3 in domestic cattle and sheep intermediate hosts in the USA and provide data for future diagnostic and epidemiological studies. Sequences have been deposited in GenBank (cox1 sequences: OR398494-OR398496, nad5 sequences: OR400695-OR400702).

Dual-RPA assay for rapid detection and differentiation of E.granulosus and E.multilocularis.

Echinococcus granulosus (Eg) and Echinococcus multilocularis (Em) are the two most widely prevalent types of echinococcosis. Several diagnostic methods have been developed for detecting Eg and Em. However, some limitations, such as being time-consuming, needing expensive instruments, or exhibiting low sensitivity, make these methods unsuitable for on-site detection. In this study, a dual-RPA assay was established to detect and differentiate Eg and Em. The primer concentration ratio, reaction time, and reaction temperature of the dual-RPA were optimized. The result showed that the primer concentration ratio of Eg:Em was 400 nM:400 nM, and the best amplification efficiency was obtained by reacting at 38 °C for 20 min. The sensitivity, specificity, and repeatability of the assay were also tested. The assay's detection limit for both Eg and Em was 10 copies/µL. The assay showed reasonable specificity by testing ten parasitic nucleic acids. The assay's intra- and inter-batch coefficients of variation were below 10%, which indicates robust reproducibility of the assay. Finally, to validate the performance of the dual-RPA assay, it was compared with real-time PCR by using 86 clinical nucleic acid samples. The coincidence rate of Eg between dual-RPA and TaqMan real-time PCR was 96.51%, and the coincidence rate of Em between dual-RPA and TaqMan real-time PCR was 98.84%, indicating its potential for accurate clinical diagnosis. Therefore, this study established a rapid and sensitive dual-RPA assay that can rapidly detect and differentiate Eg and Em in one reaction tube and provided a new assay for the detection of echinococcosis in the field.

Weighted gene co-expression network analysis reveals immune evasion related genes in Echinococcus granulosus sensu stricto.

Cystic echinococcosis (CE) is a zoonotic disease caused by the tapeworm Echinococcus granulosus sensu lato (s.l). In the intermediate host, this disease is characterized by the growth of cysts in viscera such as liver and lungs, inside of which the parasite develops to the next infective stage known as protoscoleces. There are records that the infected viscera affect the development and morphology of E. granulosus s.l. protoscolex in hosts such as buffalo or humans. However, the molecular mechanisms that drive these differences remains unknown. Weighted gene co-expression network analysis (WGCNA) using a set of RNAseq data obtained from E. granulosus sensu stricto (s.s.) protoscoleces found in liver and lung cysts reveals 34 modules in protoscoleces of liver origin, of which 12 have differential co-expression from protoscoleces of lung origin. Three of these twelve modules contain hub genes related to immune evasion: tegument antigen, tegumental protein, ubiquitin hydrolase isozyme L3, COP9 signalosome complex subunit 3, tetraspanin CD9 antigen, and the methyl-CpG-binding protein Mbd2. Also, two of the twelve modules contain only hypothetical proteins with unknown orthology, which means that there are a group of unknown function proteins co-expressed inside the protoscolex of liver CE cyst origin. This is the first evidence of gene expression differences in protoscoleces from CE cysts found in different viscera, with co-expression networks that are exclusive to protoscoleces from liver CE cyst samples. This should be considered in the control strategies of CE, as intermediate hosts can harbor CE cysts in liver, lungs, or both organs simultaneously.

Reinfection of farm dogs following praziquantel treatment in an endemic region of cystic echinococcosis in southeastern Iran.

Cystic Echinococcosis (CE) as a prevalent tapeworm infection of human and herbivorous animals worldwide, is caused by accidental ingestion of Echinococcus granulosus eggs excreted from infected dogs. CE is endemic in the Middle East and North Africa, and is considered as an important parasitic zoonosis in Iran. It is transmitted between dogs as the primary definitive host and different livestock species as the intermediate hosts. One of the most important measures for CE control is dog deworming with praziquantel. Due to the frequent reinfection of dogs, intensive deworming campaigns are critical for breaking CE transmission. Dog reinfection rate could be used as an indicator of the intensity of local CE transmission in endemic areas. However, our knowledge on the extent of reinfection in the endemic regions is poor. The purpose of the present study was to determine E. granulosus reinfection rate after praziquantel administration in a population of owned dogs in Kerman, Iran. A cohort of 150 owned dogs was recruited, with stool samples collected before praziquantel administration as a single oral dose of 5 mg/kg. The re-samplings of the owned dogs were performed at 2, 5 and 12 months following initial praziquantel administration. Stool samples were examined microscopically using Willis flotation method. Genomic DNA was extracted, and E. granulosus sensu lato-specific primers were used to PCR-amplify a 133-bp fragment of a repeat unit of the parasite genome. Survival analysis was performed using Kaplan-Meier method to calculate cumulative survival rates, which is used here to capture reinfection dynamics, and monthly incidence of infection, capturing also the spatial distribution of disease risk. Results of survival analysis showed 8, 12 and 17% total reinfection rates in 2, 5 and 12 months following initial praziquantel administration, respectively, indicating that 92, 88 and 83% of the dogs had no detectable infection in that same time periods. The monthly incidence of reinfection in total owned dog population was estimated at 1.5% (95% CI 1.0-2.1). The results showed that the prevalence of echinococcosis in owned dogs, using copro-PCR assay was 42.6%. However, using conventional microscopy, 8% of fecal samples were positive for taeniid eggs. Our results suggest that regular treatment of the dog population with praziquantel every 60 days is ideal, however the frequency of dog dosing faces major logistics and cost challenges, threatening the sustainability of control programs. Understanding the nature and extent of dog reinfection in the endemic areas is essential for successful implementation of control programs and understanding patterns of CE transmission.

Molecular characterization and haplotypes of hydatid cyst isolates collected from humans and ruminants in Setif Province (northeast of Algeria) based on mitochondrial cytochrome C oxidase subunit 1 (mt-CO1) gene sequences.

Cystic echinococcosis (CE) is a disease that can be transmitted from animals to humans, caused by the metacestode of Echinococcus granulosus. The disease has significant health and economic impacts worldwide, particularly in endemic areas. The study aimed to evaluate the prevalence of hydatid cysts in ruminants (cattle and sheep) (n = 2060) from the Setif Province of Algeria using microscopy. The results showed that hydatid cysts were detected in 9.6% (198/2060) of ruminants, with a higher prevalence in cattle (16.8%; 56/333) compared to sheep (8.2%; 142/1727). Molecular techniques were used to analyze a subset of animals consisting of 30 sheep and 4 cattle. Specifically, a fragment of the mitochondrial cytochrome c oxidase subunit 1 (mt-CO1) gene was sequenced and compared to sequences from seven humans from the same region. The results indicated that all isolates were identified as E. granulosus sensu stricto. Haplotype analysis identified 19 E. granulosus s.s. haplotypes arranged like a star, with the dominant haplotype (Hap04) at the center. Hap04 has been assigned a total of 17 positives, including positives from sheep, cattle, and two humans. This study is noteworthy for being the first to use a molecular approach to human and ruminant echinococcosis in Setif, a significant breeding region in Algeria.

Understanding the role of pigs in the transmission of zoonotic Echinococcus ortleppi in Haryana, India.

The etiological agents of zoonotic cystic echinococcosis comprise the Echinococcus granulosus sensu lato (s.l.) species complex. The present study was aimed at investigating the zoonotic genotypes of Echinococcus granulosus s.l. circulating in the pig population of Haryana, India. Out of 253 slaughtered pigs screened, 5 showed the presence of hydatid cysts. The amplification of the partial mitochondrial NADH dehydrogenase subunit 1 (nad1) gene for the molecular confirmation and phylogenetics of the retrieved metacestodes (n = 2) revealed the presence of E. ortleppi. The sequences generated herein exhibited 99.80% homology to the GenBank archived E. ortleppi sequences. Cladistics targeting genetic diversity and haplotype network analysis involved 37 E. granulosus s.l. GenBank archived sequences from India corresponding to different hosts (large and small ruminants and humans) along with the sequences (n = 2) generated in the present study. Overall, 14 haplotypes with high haplotype (0.780 ± 0.059) and low nucleotide (0.033 ± 0.010) diversities were recorded for the overall data set, which evinced a population expansion. The median-joining haplotype network revealed a stellate shape of E. granulosus sensu stricto (s.s.) sequences, which was indicative of rapid population expansion. High genetic differentiation (FST = 0.840 - 0.983) and low gene flow (Nm = 0.003 - 0.047) were recorded between the pig intermediate hosts infected with E. ortleppi and other hosts infected with E. granulosus s.s. The findings are of paramount significance for the formulation of effective control strategies considering the public health and economic impact of cystic echinococcosis.

Mouse model of secondary cystic echinococcosis.

Cystic echinococcosis (CE) is a parasitic zoonosis caused by the larval stage of the cestode Echinococcus granulosus sensu lato (s. l.), a genetic complex composed of five species: E. granulosus sensu stricto (s. s.), E. equinus, E. ortleppi, E. canadensis, and E. felidis. The parasite requires two mammalian hosts to complete its life cycle: a definitive host (mainly dogs) harboring the adult parasite in its intestines, and an intermediate host (mostly farm and wild ungulates) where hydatid cysts develop mainly in the liver and lungs. Humans are accidental intermediate hosts, being susceptible to either primary or secondary forms of CE; the first one due to the ingestion of oncospheres, and the second one because of the spillage of protoscoleces (PSC) contained within a primary cyst. Secondary CE is a serious medical problem, and can be modeled in immunocompetent mice (a non-natural intermediate host) through the intraperitoneal inoculation of viable PSC from E. granulosus s. l. This model is useful to study not only the immunobiology of CE, but also to test new chemotherapeutics or therapeutical protocols, to explore novel vaccine candidates, and to evaluate alternative diagnostic and/or follow-up tools. The mouse model of secondary CE involves two sequential stages: an early stage of parasite pre-encystment (PSC develop into hydatid cysts in the peritoneal cavity of mice), and a late or chronic stage of parasite post-encystment (already differentiated cysts slowly grow during the whole host lifespan). This model is a time-consuming infection, whose outcome depends on several factors like the parasite infective dose, the mouse strain, and the parasite species/genotype. Thus, such variables should always be adjusted according to the research objectives. Herein, the general materials and procedures needed to establish secondary CE in mice are described, as well as several useful tips and recommendations.

Assessing the Potential Apoptotic Effects of Different Hydatid Cyst Fluids on Human Healthy Hepatocytes and Hepatocellular Carcinoma Cells.

Cystic Echinococcosis (CE) is a zoonotic infection caused by the larval form of Echinococcus granulosus in humans. Emerging evidence suggests an intriguing inverse association between E. granulosus infection and the occurrence of cancer. This study aimed to investigate the influence of diverse host-derived hydatid cyst fluids (HCF) with distinct genotypes on human liver hepatocytes (HC) and hepatocellular carcinoma cells (HepG2). Specifically, we examined their effects on cell proliferation, apoptosis sensitivity (BAX/BCL-2), apoptosis-related p53 expression, and the expression of cancer-related microRNA (hsa-miR-181b-3p). Cell proliferation assays, real-time PCR, and ELISA studies were conducted to evaluate potential anti-cancer properties. The findings revealed that animal-origin HCF (G1(A)) induced direct cell death by augmenting the susceptibility of HepG2 cells to apoptosis. Treatment with both G1(A) and G1(H) HCF sensitized HepG2 and HC cell lines to apoptosis by modulating the BAX/BCL-2 ratio, accompanied by upregulation of the p53 gene. Additionally, G1(A) HCF and human-derived HCFs (G1(H), G7(H)) reduced the expression of miR-181b-3p in HepG2 cells. Consequently, this study demonstrates the potential anti-cancer effect of HCF in HepG2 cells and provides the first comparative assessment of HCFs from human and animal sources with diverse genotypes, offering novel insights into this field.

Echinococcus granulosus sheep strain (G1) as the predominant genotype in definitive host (dogs) isolates in northeastern Iran.

Echinococcus granulosus sensu lato (E. granulosus s.l.) is a zoonotic parasite, causing cystic echinococcosis in humans. In the present study, prevalence and genotypes of E. granulosus s.l. was assessed in stools collected from 244 dogs including 138 stray and 106 domestic animals using high resolution melting curve (HRM) method. Initially, to detect taeniid eggs in feces, all samples were examined using the formalin-ether techniques. Genomic DNA was extracted from the positive samples and E. granulosus s.l. was differentiated from other Taeniidae parasites using SSU-rDNA gene and E. granulosus s.l. was analyzed for genotyping using HRM based on the cox1 gene. In total, 12.7% (31/244) of the samples were positive for Taeniidae eggs. In addition, among the positive samples, 77.4% (24/31) were positive for E. granulosus s.l.. In details, 11.3% (12/106) of the domestic dogs and 8.7% (12/138) of the stray dogs were positive for E. granulosus s.l.. The results of HRM analysis showed that all E. granulosus s.l. isolates were G1 strain. Findings of the present study indicated a considerable prevalence of E. granulosus G1 among dogs in the northeast of Iran and imply a serious risk of transmitting to humans and livestock.

G1 the common Echinococcus granulosus genotype infected domestic cat (Felis catus) in Iraq.

BACKGROUND: Infections of cats with Echinococcus granulosus is uncommon because the cat is not part of the parasite life cycle that a carnivorous and another herbivore represent. Nevertheless, it occurs incidentally when eating food or drinking water contaminated with the worm's larva, especially with the presence of the definitive host (dogs), in this case, the infections are concentrated in stray or outside cats. For this reason, this study examined the possibility of cat infection with E. granulosus and diagnosed the common genotype of this infection. OBJECTIVE: This study examined the possibility of cat infection with E. granulosus and diagnosed the common genotype of this infection. METHODS: Four of the 37 cats that had died in different accidents developed cystic echinococcosis (CE). The cytochrome c oxidase subunit I (COX1) gene was initially amplified and sequenced to determine if these cysts belonged to E. granulosus, in beginning. The DNA fragments resulting from sequencing were then compared and aligned with other sequences using the Gene Bank database. Finally, a phylogenetic tree was drawn according to the sequence data obtained from cox1 genes sequencing, and the MEGA 7.0 phylogenetic analysis program was utilized. RESULTS: Four different sequences were deposited in the Gen Bank with accession numbers (ON795961 to ON795964), all of which belong to the G1 genotype. Approximately 84% and 100% of these sequences aligned with G1 (AB622277.1) and G1 (MG722980.1), respectively. CONCLUSIONS: G1 is the dominant genotype that causes cat infections, even though the cat's EC infection was incidental.