Parasite-derived microRNA let-7-5p detection for metacestodiasis based on rolling circular amplification-assisted CRISPR/Cas9.

Metacestodiasis is an infectious disease caused by the larval stage of cestode parasites. This disease poses a serious health hazard to wildlife, livestock, and humans, and it incurs substantial economic losses by impacting the safety of the livestock industry, the quality of meat production, and public health security. Unfortunately, there is currently no available molecular diagnostic method capable of distinguishing cysticercus- and Echinococcus-derived microRNAs (miRNAs) from other helminthes and hosts in the plasma of metacestode-infected animals. This study aims to develop a specific, sensitive, and cost-efficient molecular diagnostic method for cysticercosis and echinococcosis, particularly for early detection. The study developed a rolling circular amplification (RCA)-assisted CRISPR/Cas9 detection method based on parasite-derived miRNA let-7-5p. Using a series of dilutions of the let-7 standard, the limit of detection (LOD) of the qPCR, RCA, and RCA-assisted CRISPR/Cas9 methods was compared. The specificity of qPCR and CRISPR/Cas9 was evaluated using four artificially synthesized let-7 standards from different species. A total of 151 plasma samples were used to evaluate the diagnostic performance. Additionally, the study also assessed the correlation between plasma levels of let-7-5p, the number of Taenia pisiformis cysticerci, and the weight of Echinococcus multilocularis cysts. The results demonstrated that the RCA-assisted CRISPR/Cas9 assay could significantly distinguish let-7 from cestodes and other species, achieving a LOD of 10 aM; the diagnostic sensitivity and specificity for rabbit cysticercosis and mouse E. multilocularis were 100% and 97.67%, and 100% and 100%, respectively. Notably, let-7-5p gradually increased in the plasma of T. pisiformis-infected rabbits from 15 days post infection (dpi), peaked at 60 dpi, and persisted until 120 dpi. In E. multilocularis-infected mice, let-7-5p gradually increased from 15 dpi and persisted until 90 dpi. Furthermore, the expression of let-7-5p positively correlated with the number of cysticerci and cyst weight. These results indicated that the let-7-5p-based RCA-assisted CRISPR/Cas9 assay is a sensitive and specific detection method that can be used as a universal diagnostic method for metacestodiasis, particularly for early diagnosis (15 dpi).

Cardiac cystic echinococcosis-A systematic review and analysis of the literature.

Human cystic echinococcosis (CE) is a parasitic infection caused by the larval stage of the tapeworm Echinococcus granulosus sensu lato, primarily affecting the liver and lungs. Although the heart is affected in only 0.02-2% of all CE cases, a considerable number of cases have been, and continue to be, published. However, due to the rare occurrence of cardiac CE and the resulting lack of clinical trials, knowledge about various aspects of the disease remains limited. To obtain a clearer picture of anatomical, clinical, diagnostic as well as therapeutic aspects of cardiac CE, we systematically reviewed the literature published between 1965 and 2022. The anatomical pattern of the affected cardiac structures follows the extension of the supplying capillary bed. The majority of patients (82.7%) are symptomatic and present with prolonged non-specific symptoms such as dyspnoea, chest pain and palpitations. Acute complications generally derive from cyst rupture, occur in 18.3% of cases and manifest as embolism, pericardial tamponade, or anaphylactic reaction in 83.2%, 17.8% and 10.9% of these cases, respectively. As for CE cysts localized in other organs, the diagnosis of cardiac CE is made by imaging. Serology plays a minor role due to its limited sensitivity. Unlike abdominal CE cysts, cardiac CE cysts are usually resected independent of their stage (active/inactive), because their presence impairs cardiac performance and carries the risk of long-term sequelae. More than 80% of patients are treated with a single surgical intervention. We found a disease-related case fatality rate of 11.1%. Since local recurrence was reported up to 108 months and secondary CE up to 72 months after surgery, patients should be followed up for a minimum of 10 years.

Exploring rare locations of hydatid disease: a retrospective case series.

INTRODUCTION: Hydatid disease, endemic in Mediterranean countries, primarily affects the liver, but can manifest in diverse organs. Non-hepatic and non-pulmonary cysts often pose diagnostic challenges. This study examines patients with hydatid cysts in atypical locations. METHODS AND RESULTS: From 2013 to 2020, our center treated 250 echinococcosis patients, among whom 11 cases (4.4%) with hydatid disease in uncommon sites were retrospectively reviewed. The distribution of unusual cyst locations and their clinical implications are discussed. CONCLUSION: Diagnosing hydatid cysts in uncommon locations is a formidable challenge. Surgeons should always contemplate the prospect of an unconventional cyst location when encountering patients with cystic masses in endemic regions. Failing to consider this possibility could lead to unfavorable outcomes.

Case Report of a Hidden Intruder-Extremely Rare Presentation of Hydatidosis as a Nuchal Tumoural Mass.

Introduction: The parasitic tapeworm impersonated by the larvae of Echinococcus granulosus represents the aetiology of the hydatid pathology. The predilect site of invasion is the liver, but there are other cases of different localization all over the body, regardless of the type of invaded tissue. Soft tissue hydatidosis can be a real challenge for the clinician in terms of the diagnosis, and it might generate various complications such as anaphylactic shock. The aim of the present work is to illustrate a unique case of primary hydatidosis located in the nuchal region. Case Report: We report the case of a 68-year-old male patient, a zootechnic, who presented at the hospital with a tumoural mass (dimension: about 12/10 cm) located in the nuchal region. The complex approach needed consisted of surgical therapy along with histopathological confirmation of the diagnostic and antiparasitic medication, which led to a complete recovery with a low probability of recurrence. Discussion: Encounters with patients with primary soft tissue hydatidosis are exceptionally rare, but the surgeon must take into consideration this clinical diagnosis, especially for patients located in an endemic region with occupations that might have exposed the patient to this type of parasite.

Detection of circulatory E. granulosus-derived cell-free DNA in the plasma and urine of human cystic echinococcosis using an in-house PCR: a potential promising diagnostic biomarker.

BACKGROUND: The diagnostic tool for identifying cystic echinococcosis (CE) patients at an early stage is currently lacking. However, circulatory cell-free DNA (cfDNA) has shown potential as a biomarker for parasitic infections and could be used for diagnosing CE. RESEARCH DESIGN AND METHODS: The plasma and urine samples were collected from 39 patients with confirmed CE through imaging and histopathological techniques. All plasma samples were tested for anti-echinococcal antibodies using a commercial ELISA test. Total plasma and urine cfDNA were extracted and an in-house PCR assay was developed to detect E. granulosus specific cfDNA in the samples of CE patients. RESULTS: Out of the 39 patients, 30 tested positive for E. granulosus using serology, with a sensitivity of 76.9%. Moreover, the detection rates for the cfDNA were 79.5% in plasma samples and 58.97% in urine samples using the 80 bp COX1 gene. The plasma-based PCR and serology test showed the highest agreement (Kappa = 0.53). CONCLUSIONS: Plasma-based PCR has been found to be a reliable diagnostic tool for identifying CE patients at different cyst stages. It offers validity, speed, and sufficient sensitivity, making it an alternative to serology in diagnosing CE in endemic areas.

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.

Concomitant Polymyalgia Rheumatica and Giant Cell Arteritis Associated with Cystic Echinococcosis: A Rare Geriatric Case.

INTRODUCTION: Parasitic infections could be an important triggering factor for autoimmune diseases. We present a clinical case of concomitant polymyalgia rheumatica (PMR) and giant cell arteritis (GCA) induced with cystic echinococcosis (CE). CASE PRESENTATION: A 74-year-old male was admitted with a 2-month history of progressive pain at the shoulders and hip, movement restriction, and constitutional symptoms. As a result of the examinations performed due to rheumatological complaints, PMR and GCA were diagnosed. The cystic appearance, which was incidentally detected in the liver 6 months ago and not examined at that time, was found to be hydatid cyst. Medical treatment was initiated for all three conditions and the patient's symptoms improved significantly. DISCUSSION: Parasite infections may cause various autoimmune diseases because of molecular mimicry or sustained immune activation. Echinococcus granulosus is a very complex multicellular parasite and highly immunogenic for humans. Some body parts of the parasite, the outer surface and secreted particles, stimulate the host immune system strongly. CONCLUSION: The first case in the literature of coexistence of PMR and GCA associated with CE. Autoimmune diseases should be evaluated in patients with CE. Furthermore, CE should be considered in patients with autoimmune diseases in the presence of a cyst.

Primary seronegative intermuscular hydatidosis in a child, an infection that should be approached like a tumor: a case report and review of the literature.

INTRODUCTION: Intermuscular hydatid cyst is one of the rarest types of hydatid cyst, and as far as we know, only nine cases were reported in the literature before this study. CASE PRESENTATION: We present a 10-year-old Iranian child with an intermuscular cystic mass in the medial-distal thigh. Despite the typical imaging findings, the patient's serological and hematological tests were negative for hydatid cyst. The cyst underwent wide excision accompanied by neoadjuvant and adjuvant chemotherapy with Albendazole. No evidence of recurrence was detected during the one-year follow-up. CONCLUSION: Hydatid cysts should always be considered in the differential diagnosis of soft tissue cystic masses in endemic areas, and aspiration or drainage should be avoided as much as possible, even when serological tests are negative and imaging is non-diagnostic. In cases where the diagnosis of a hydatid cyst has been confirmed before the surgery, it is recommended to approach the cyst, like a tumor with chemotherapy using Albendazole both before and after wide cyst excision.

Investigation of patients with new infection of echinococcal cyst in Sardinia, Italy.

Cystic Echinococcosis (CE) is a zoonotic disease caused by the larval stage of a tapeworm of Taeniidae family, genus Echinococcus and species Echinococcus granulosus sensu lato (s.l.). CE is a worldwide public health problem and is highly incident in all Mediterranean areas. Following clinical, image techniques and serological investigations all 83 subjects involved in the study were diagnosed for CE. General and clinical data were entered into a database and evaluated. The 43.37% were female and 56.63% male, mean age was 50.71 while the range most represented (22.7%) was between 61->70 years. The purposes of our survey were to investigate these 83 patients enrolled in the study and to deeply examine 20 (24.10%) patients that developed a new echinococcal cyst. Moreover, the causes at the basis of the onset of a new cyst were analysed, together with a possible correlation with different treatments related to primary infection corresponding to surgery (n=7), albendazole (n=6), PAIR (n=3) and watch and wait (n=4). A possible link with medical treatments of the primary infection was observed in the subjects who underwent surgery or PAIR and a likely correlation attributable to high environmental contamination in subjects managed with drugs or watch and wait was detected. Moreover, our analysis evidenced that patients with a new infection presented a more severe diagnosis along with a major mortality rate. Finally, these data may have an important contribution for an epidemiological point of view concerning the percentage of CE in a specific geographical endemic area, such as Sardinia.

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.

Additional diagnoses of Echinococcus multilocularis in eastern chipmunks (Tamias striatus) from southern Ontario - results from ongoing surveillance for E. multilocularis in intermediate hosts in Ontario, Canada.

Echinococcus multilocularis, a cestode with zoonotic potential, is now known to have a high prevalence in wild canid definitive hosts of southern Ontario. The distribution of E. multilocularis across this region in red foxes (Vulpes vulpes) and coyotes (Canis latrans) is widespread yet heterogenous. In contrast, confirmed diagnoses of E. multilocularis in wild free-ranging intermediate hosts within Ontario are currently limited to a single eastern chipmunk (Tamias striatus). These findings prompted ongoing surveillance efforts in intermediate host species, primarily rodents. Our report describes the results of passive surveillance through wildlife carcass submissions to the Canadian Wildlife Health Cooperative (CWHC) and targeted active sampling of small mammal species from 2018 to 2023; a second and third eastern chipmunk were found to be infected with E. multilocularis. However, these were the only occurrences from surveillance efforts which collectively totaled 510 rodents and other small mammals. Continued surveillance for E. multilocularis in intermediate hosts is of high importance in light of the recent emergence of this parasite in Ontario.

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.

Diagnosis and treatment experience of atypical hepatic cystic echinococcosis type 1 at a tertiary center in China.

BACKGROUND: Some hydatid cysts of cystic echinococcosis type 1 (CE1) lack well-defined cyst walls or distinctive endocysts, making them difficult to differentiate from simple hepatic cysts. AIM: To investigate the diagnostic methods for atypical hepatic CE1 and the clinical efficacy of laparoscopic surgeries. METHODS: The clinical data of 93 patients who had a history of visiting endemic areas of CE and were diagnosed with cystic liver lesions for the first time at the People's Hospital of Xinjiang Uygur Autonomous Region (China) from January 2018 to September 2023 were retrospectively analyzed. Clinical diagnoses were made based on findings from serum immunoglobulin tests for echinococcosis, routine abdominal ultrasound, high-frequency ultrasound, abdominal computed tomography (CT) scan, and laparoscopy. Subsequent to the treatments, these patients underwent reexaminations at the outpatient clinic until October 2023. The evaluations included the diagnostic precision of diverse examinations, the efficacy of surgical approaches, and the incidence of CE recurrence. RESULTS: All 93 patients were diagnosed with simple hepatic cysts by conventional abdominal ultrasound and abdominal CT scan. Among them, 16 patients were preoperatively diagnosed with atypical CE1, and 77 were diagnosed with simple hepatic cysts by high-frequency ultrasound. All the 16 patients preoperatively diagnosed with atypical CE1 underwent laparoscopy, of whom 14 patients were intraoperatively confirmed to have CE1, which was consistent with the postoperative pathological diagnosis, one patient was diagnosed with a mesothelial cyst of the liver, and the other was diagnosed with a hepatic cyst combined with local infection. Among the 77 patients who were preoperatively diagnosed with simple hepatic cysts, 4 received aspiration sclerotherapy of hepatic cysts, and 19 received laparoscopic fenestration. These patients were intraoperatively diagnosed with simple hepatic cysts. During the follow-up period, none of the 14 patients with CE1 experienced recurrence or implantation of hydatid scolices. One of the 77 patients was finally confirmed to have CE complicated with implantation to the right intercostal space. CONCLUSION: Abdominal high-frequency ultrasound can detect CE1 hydatid cysts. The laparoscopic technique serves as a more effective diagnostic and therapeutic tool for CE.

Primary hydatid cyst of the spleen: A rare case report and literature review.

Hydatidosis is an endemic disease in certain areas in the world particularly in the Mediterranean, the Middle East, and South America, caused by a cestode known as Echinococcus granulosus. Humans are the accidental intermediate hosts. The liver and the lungs are the most commonly involved organ. If the parasite passes through the pulmonary capillary bed, the hydatid cyst may develop at any site in the body like bone, pancreas, brain, kidney, and orbit. Isolated spleen hydatid cyst is very rare. We hereby report one observation of isolated hydatid cyst of the spleen in a patient living in non-endemic area and without any potential risk.

Gossypiboma mimicking hepatic hydatid cyst after 22 years: a case report.

Gossypiboma is a non-absorbable material that is forgotten during surgery. These are medicolegal pathology that leads to diagnostic confusion from abscess to tumor. We present the case of gossypiboma detected in laparotomy in a 57-year-old male patient who had a history of operation due to a hydatid cyst 22 years ago and was referred to our center due to hydatid recurrence. The case should suggest a history of surgical gossypiboma, especially in asymptomatic patients. In this preventable pathology, the strategy during sur-gery and the careful and proper attitude of the surgical team are the main determinants.

Confirmed Autochthonous Case of Human Alveolar Echinococcosis, Italy, 2023.

In September 2023, a patient in Italy who had never traveled abroad was referred for testing for suspected hepatic cystic echinococcosis. Lesions were incompatible with cystic echinococcosis; instead, autochthonous alveolar echinococcosis was confirmed. Alveolar echinococcosis can be fatal, and awareness must be raised of the infection's expanding distribution.

A Very Rare Cause of Dyspnea in a Child: A Hydatid Cyst From Echinococcus.

Hydatid cyst is a human parasitic disease caused by echinococcus granulosus that affects mainly the liver or the lungs but may be found in any organ, including the heart in up to 2% of the cases. Humans are infected as accidental hosts by contaminated vegetables or water and by contact with saliva from infected animals. Although cardiac echinococcosis can be fatal, it is rare and often asymptomatic in the early stage. We present the case of a young boy living on a farm who suffered from mild exertional dyspnea. He suffered from pulmonary and cardiac echinococcosis and was treated surgically through median sternotomy to prevent potential cystic rupture.