Strobilocercus fasciolaris induced hepatic sarcoma in Wistar Rats

Strobilocercus fasciolaris induced hepatic sarcoma in Wistar Rats | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Strobilocercus fasciolaris induced hepatic sarcoma in Wistar Rats Rahul Kumar, Vikrant Sudan This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-4588055/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Background Taenia taeniaeformis infects the small intestine of domestic and wild carnivores. Rodents, the intermediate hosts, ingest the gravid proglottids from the carnivores' feces. The parasite’s oncosphere reaches the liver via portal circulation after penetrating the wall of small intestine. Purpose The present study reports hepatic sarcoma caused by strobilocercus- the metacestodal stage of Taenia taeniformis in laboratory rats. Method Twenty-three rats in a colony of 72 male wistar rats died suddenly with clinical signs of weakness, lethargy and mild diarrhea. The carcasses were sent to the department of veterinary pathology for postmortem examination. Results Grossly, the liver of 18 rats were having multiple, cream to white cysts of variable sizes of Strobilocercus fasciolaris . Histopathologically, the hepatic tissue surrounding the cysts revealed zones of fatty change, inflammation with infiltration of plasma cells, macrophages and eosinophils, presence of granulation tissue, metaplasia and fibrosarcoma. The pathological changes in the stomach and small intestines include hyperplasia of gastric and intestinal glands and their mucosa. Conclusion The likelihood of altered research results of studies conducted in immune-compromised T. taeniformis infected rats and its zoonotic implications are discussed herewith. Hepatic sarcoma Strobilocercus fasciolaris Wistar rats Histopathology Figures Figure 1 Figure 2 1. Introduction Laboratory animals, especially rodents such as rabbits, guinea pigs, mice, and rats, are widely utilized as models for investigating the pathobiology of various microbial or toxicological agents. The outcomes of these experimental studies can be significantly influenced by both biotic and abiotic factors, including environmental conditions, nutrition, and infection status [11, 27, 30]. Parasitic infections, in particular, can affect research results by altering the physiological and immunological characteristics of the hosts, potentially modifying their susceptibility to experimental stress. These alterations may lead to tissue damage, abnormal tissue growth, nutrient competition, blood volume reduction, and mechanical interference [15, 27]. Taenia taeniaeformis, though globally widespread, is rarely found in laboratory animals [29]. This parasite primarily infects wild and domestic carnivores, which serve as its definitive hosts. Rodents, such as rats, act as intermediate hosts, becoming infected by ingesting feed or water contaminated with the eggs from definitive hosts. Once ingested, the eggs hatch in the small intestine of the rodent, and the resulting embryos travel via the portal circulation to the liver, where they develop into the infective larval stage, known as Strobilocercus fasciolaris or Cysticercus fasciolaris , or simply strobilocercus [32, 5]. Felines and other carnivorous definitive hosts become infected by consuming the liver of rats harboring the larval stage [29]. Another source of rodent infection is during the transportation of rats between the source and host institutions. Transportation stress often exacerbates these latent infections. As laboratory animal facilities often lack proper screening and quarantine measures for parasitic infestations, these infections can easily spread throughout the colony, infecting other animals . The pathology of strobilocercus-induced liver disease in rodents begins with the ingestion of T. taeniaeformis eggs, which develop into larvae that form cysts in the liver parenchyma of rats, leading to progressive chronic inflammation and vigorous fibroplasia [6, 31]. Strobilocercus lodges in the liver parenchyma [6, 31], causing physiological and immunological changes, tissue damage, abnormal growth, nutrient competition, and deviations in animal health and research outcomes [15]. Additionally, fibroplastic and inflammatory lesions may progress to fibrosarcoma [20, 24]. There have been reports of the spontaneous occurrence of Taenid cyst-associated hepatic mesenchymal tumors with rare systemic metastasis and gastroenteropathy [10, 14, 20]. The parasite also poses a zoonotic risk to animal house workers, as humans can act as accidental hosts, with occasional reports of the parasite in the intestines and liver of infected individuals [12, 13, 36, 27]. This study reports the incidental findings and pathology of Strobilocercus in laboratory rats, including the rare occurrence of associated hepatic sarcoma. Additionally, the study examines the potential impact on research outcomes, particularly in immunocompromised rats, and discusses its zoonotic implications. 2. Materials and methods 2.1. Ethical Approval Three-month-old Wistar rats were maintained in the university's laboratory animal house facility for another approved study (IAEC approval no. 110/IAEC/16). The animals were housed according to the guidelines for the care and use of animals in scientific research, as specified by the Institutional Animal Ethical Committee registered with the Committee for the Purpose of Control and Supervision of Experimental Animals (CPCSEA). During the study period, 23 rats died over the course of two weeks, displaying clinical signs of mild diarrhea, lethargy, and anorexia. 2.2. Necropsy and sample collection A meticulous and scientific necropsy was performed on the deceased rats. All visceral organs were thoroughly inspected for the presence of cysts, adult parasites, or other gross lesions. For microscopic investigations, representative tissue samples containing parasitic cysts in situ, as well as some parasites, were carefully preserved in 10% neutral buffered formalin. All significant findings were documented with photographs. 2.3. Morphological studies The liver samples, harboring parasitic cysts, were carefully isolated and submerged in a Petri dish filled with normal saline. The morphology of the developmental stages was then scrutinized for identification, involving the delicate dissection of the outer cyst covering from freshly collected livers [33]. A comprehensive gross morphological evaluation was carried out to ascertain the average number and size of the cysts. Subsequently, the cysts underwent precise dissection to assess the presence or absence of larval stages. 2.4. Staining of developmental stages The liver lobes containing the cysts were washed in phosphate-buffered saline (1% PBS). The cyst wall was carefully dissected to collect the metacestode larva. The larva was then pressed between two glass slides and fixed in 10% neutral buffered formalin for 72-96 hours [7]. It was subsequently washed overnight in running tap water, stained with borax carmine, treated with acid alcohol, cleared in xylene, and finally mounted in dextrin plasticized xylene (DPX) (Meyer and Olsen, 1975). The development of scolices and the complete arrangement of the rostellar hooks and suckers were studied according to the method described by Meyer and Olsen (1975). Additionally, the larva was stained with H&E stain using the paraffin embedding technique for histopathology [23]. 2.5. Histopathology Representative tissue samples from livers containing cysts, as well as intestines and individual parasites, were collected and fixed in 10% neutral buffered formalin. The developmental stages of the parasites and the tissues were then sectioned and processed for histopathological examination [23]. 3. Results Twenty-three deceased Wistar rats, aged 3 months, were submitted to the Department of Veterinary Pathology for necropsy and disease diagnosis. Grossly, cream-colored parasitic cysts were observed embedded in the hepatic parenchyma of 18 rats (78%) (Fig. 1A, 1B). On cut section, the metacestode larva was found inside the cysts (Fig. Inset 1B). Some rats had multiple parasitic cysts embedded in the liver parenchyma (Fig. 1B), containing parasites approximately 4 inches in length. 3.1. Parasite and Larval Morphology The livers of the rats exhibited multiple cream-colored cysts affecting single or multiple lobes (Fig. 1A and 1B), ranging in size from 3-5mm. The recovered developmental stages measured between 4-20 inches in length, with relatively larger scolexes measuring 1-2 inches in length and a long neck (Fig. inset 1B). The larval body displayed pseudo segmentation throughout its length with a terminal bulged portion (Fig. Inset 1B). Stained parasites exhibited typical taenid morphology. In the larger cysts, well-developed scolices were observed (Fig. 1C), while most cysts were smaller in size. The larger scolices possessed two distinct rows of characteristic penknife-shaped hooks (Fig. 1C and 1D). A total of 42 hooks, consisting of 21 large and 21 small hooks, were recorded (Fig. Inset 1D). The mean size of the larger hooks was 390.89 ± 10.08 μm, and that of the smaller hooks was 241.46 ± 14.13 μm. 3.2. Histopathology Microscopic examination of the affected liver sections revealed vacuolation, fatty changes, and inflammatory alterations with extensive infiltration of a mixed cellular population including lymphocytes, macrophages, and eosinophils (Fig. 2A and 2B). The hepatic tissue surrounding the cyst exhibited metaplastic changes and a proliferated fibrous connective tissue layer with strobilocercus within the cyst lumen (Fig. 2A and 2B). The cyst wall displayed a thin connective tissue capsule containing mature larger developmental stages and a thick wall of connective tissue in comparatively younger juvenile developmental stages. Furthermore, a zone of granulation tissue with lymphocytic cuffing was observed around certain cysts (Fig. 2A (black arrow) and 2B (black arrow)). Some liver sections showed hepatic parenchymal changes suggestive of neoplastic transformation of sarcoma (Fig. 2C and 2D). These neoplastic changes included poorly defined interwoven bundles of spindle cells that were pleomorphic, hyperchromatic, with bizarre nuclei, abundant eosinophilic cytoplasm, and separated by collagen as intercellular matrix (Fig. 2C and 2D). Infiltration of plasma cells, macrophages, and eosinophils surrounding the infected areas was also observed. Some sections suggestive of hepatic sarcomas exhibited tumor giant cells and 2-8 mitotic figures per high-power microscopic field (Fig. 2C (black arrow) and 2D (black arrow)). Hepatocytes in unaffected zones and at a distance from the cysts showed mild fatty changes and apparently healthy liver tissue. Hyperplasia of gastric mucosa, distension of gastric glands, and a marked increase in the number of goblet cells in the duodenal mucosa were also noted. No histopathological changes were observed in any other visceral organs. 4. Discussion Taenia taeniaeformis parasitizes the small intestine of both domestic and wild carnivores. Intermediate hosts, such as rodents, ingest the gravid proglottids passed in the feces of the definitive hosts. The parasite oncosphere traverses the stomach of the intermediate host and is released in their small intestine, penetrating the intestinal wall via the portal circulation and then carried to the liver, where it lodges into hepatic capillaries. Approximately six days post-infection, they are found inside small vesicles formed by the proliferation of host connective tissue on the liver surface. After a maturation period of 6-8 months, the developmental stages become fully grown infective strobilocerci with caudal vesicles. This parasite is cosmopolitan, with reports from various regions including India [20, 34], Japan [25], Egypt [35], and Mexico [29]. Experimental studies have revealed a relationship between the implantation of developmental stages and the development of liver tumors [8]. In this study, Wistar rats exhibited signs of weight loss, lethargy, and mild diarrhea before succumbing. Similar clinical signs of gastroenteropathy have been documented previously [20, 16, 34]. Hepatic tumors induced by T. taeniaeformis led to nonspecific clinical signs such as lethargy, weight loss, anorexia, and sudden death [14]. Infected rats displayed nonspecific clinicopathological changes that could significantly affect the outcomes of experimental studies. These changes may include elevated activity of liver-specific enzymes, decreased concentration of glucose, and an increase in neutrophils, lymphocytes, and eosinophils count in peripheral blood smears [4, 18]. Of the 23 rats submitted to the Department of Veterinary Pathology for necropsy, eighteen rats (18/23) were found to have cystic liver i.e., an incidence of 78%. The laboratory animal house maintains hygienic conditions and adheres to strict biosecurity measures; hence, the possible sources of infection could be the breeder colony or potential breaches in biosecurity protocols during transportation between the source and host institution. All rats were 3 months old, yet they exhibited cysts of varying sizes (3-5mm) and maturity levels. Gross morphological examination of the cysts revealed variability in number, size, and maturity, likely due to differences in exposure time and age at the time of infection. Another potential factor could be the variance in the immune response of individual rats to the parasite. Hanes [14] noted that rats with infections less than one month old typically had few immature cysts without developed scolices, affecting only 1-2 hepatic lobes. Conversely, rats with infections exceeding one month tended to have a higher number of cysts affecting the liver parenchyma more severely and involving a greater number of hepatic lobes. The scolex consistently contained two rows of characteristic penknife-shaped hooks with an equal number of large and small hooks in all cases. However, some researchers [2] have reported 30–40 hooks in the rostellum of the metacestode. Microscopic examination of affected liver sections revealed fatty, inflammatory, metaplastic changes in some areas, and proliferative fibrous tissue changes in the liver parenchyma. The infiltration of mononuclear cells in the hepatic parenchyma surrounding the cyst observed in this study aligns with previous reports [30, 20, 3, 22]. Histopathological findings were indicative of fibrosarcomatous changes characterized by spindle-shaped pleomorphic neoplastic cells in the liver parenchyma. Similar lesions have been reported in Sprague-Dawley rats [14, 20, 16]. Chronic infestation of larvae in the liver parenchyma induces chronic irritation and inflammatory changes, which can progress to metaplasia and ultimately neoplasia [1, 16]. The specific pathogenesis of neoplastic changes in the liver is not fully understood, but a causal association of T. taeniaeformis with hepatic tumors is well established [20, 16]. The causal hypothesis suggests the involvement of the parasite, larval-derived oncogenic substances, chronic inflammation, irritation of liver parenchyma due to the presence of larval cysts, oxidative damage, and immunosuppression caused by the adult parasite and larvae themselves [8, 9, 14]. Histopathological changes in the gastrointestinal tract (GIT) include hyperplasia of mucosal glands and hypertrophy of the mucosa. Infected rats may develop lesions of inhibited gastric acid secretion, hypergastrinemia, and gastric and intestinal mucosal hyperplasia [18, 26]. These changes are attributed to larval secretions in the gastric and intestinal lumen [21]. 5. Conclusion The current study documents Strobilocercus fasciolaris infection in Wistar rats, leading to the subsequent development of hepatic sarcoma. The developmental stage of the parasite acts as a foreign body, causing gastroenteropathy and chronic irritation and inflammation of the liver parenchyma, ultimately resulting in neoplasia. This report underscores the importance of screening laboratory animals for parasitic infestations and implementing quarantine measures before introducing them into the colony. Additionally, this infection presents a small but significant zoonotic risk to laboratory animal house workers and animal handlers. Infection can occur through contamination of clothing and other personal items while handling infected animals, their feed, feeders, utensils, bedding materials, etc., or due to breaches in biosecurity protocols. Declarations Acknowledgments Authors are thankful to Head, Department of Veterinary Microbiology, Department of Veterinary Pathology and Dean, College of Veterinary Science and Animal Husbandry for providing the facilities to conduct this study. Author’s contribution RK performed the histopathological examination and wrote the manuscript. VK identified the parasite and edited the final draft. Source of funding Not applicable. Data accessibility statement Not applicable. Ethical statement The study was ethically approved by Institutional Animal Ethical Committee (IAEC) approval no. 110/IAEC/16. Consent for publication Not applicable. Conflict of interest None of the authors is having conflict of interest. References Al-Jashamy, K., El Salihi, K., Sheikh, A. and Saied, H., 2004, May. Cysticercosis in rat infected with C. fasciolaris. In Proceedings of 9th National conference on medical sciences. University Sciences Malaysia, Kubang Kerian (p. 185). Al-Jashamy, K. and Islam, M.N., 2007. 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A parasitological survey of zoonotic cestodes carried by house rats in Aswan, Egypt, reveals cryptic diversity at the molecular level. Veterinary World , 14 (8), p.2160. Zibaei, Mohammad, Seyed Mahmoud Sadjjadi, and Bahador Sarkari., 2007. Prevalence of Toxocara cati and other intestinal helminths in stray cats in Shiraz, Iran. Trop Biomed, 24 (2), 39-43. Additional Declarations No competing interests reported. Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-4588055","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":321903411,"identity":"d8cb59b6-90e1-4565-acac-5748b1914f28","order_by":0,"name":"Rahul Kumar","email":"data:image/png;base64,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","orcid":"","institution":"Uttar Pradesh Pandit Deen Dayal Upadhyaya Pashu Chikitsa Vigyan Vishwavidyalaya Evam Go-Anusandhan Sansthan","correspondingAuthor":true,"prefix":"","firstName":"Rahul","middleName":"","lastName":"Kumar","suffix":""},{"id":321903412,"identity":"42666ce1-21e1-4f22-a317-f116cdef21d8","order_by":1,"name":"Vikrant Sudan","email":"","orcid":"","institution":"Guru Angad Dev Veterinary and Animal Sciences University (GADVASU)","correspondingAuthor":false,"prefix":"","firstName":"Vikrant","middleName":"","lastName":"Sudan","suffix":""}],"badges":[],"createdAt":"2024-06-16 01:38:16","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-4588055/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-4588055/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":59546910,"identity":"1948ee40-2354-4e60-91cc-c1de5d6f2943","added_by":"auto","created_at":"2024-07-03 05:21:14","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":392639,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eA) \u003c/strong\u003eOpen abdomen of a dead rat showing liver with white colored \u003cem\u003eStrobilocercus fasciolaris\u003c/em\u003e cysts. B) \u003cem\u003eStrobilocercus fasciolaris\u003c/em\u003e larval cyst embedded in the liver parenchyma. Inset B\u003cem\u003e Strobilocercus fasciolaris\u003c/em\u003elarva with cyst capsule, large scolex, long neck and pseudo segmentation of entire body length with terminal bulged portion. C) Mature scolex of \u003cem\u003eStrobilocercus fasciolaris\u003c/em\u003e D \u0026amp; Inset D) Higher magnification of scolex with two distinct rows of large and small hooks.\u003c/p\u003e","description":"","filename":"1.png","url":"https://assets-eu.researchsquare.com/files/rs-4588055/v1/aacfa8cd589e85edf7679211.png"},{"id":59546911,"identity":"6ebe9248-0d93-472c-9706-96a73a4b7154","added_by":"auto","created_at":"2024-07-03 05:21:14","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":495874,"visible":true,"origin":"","legend":"\u003cp\u003eHistopathology of parasitized liver: (A) Section of liver showing \u003cem\u003eCysticercus fasciolaris\u003c/em\u003elarva invaded deeply in the liver parenchyma and infiltration of lymphocytes, plasma cells and a few eosinophils. (X100 H\u0026amp;E). B) Section of liver of Rat showing thick capsule with mononuclear cell Infiltration around a \u003cem\u003eCysticercus fasciolaris\u003c/em\u003e surrounded by spindle shaped cells forming interwoven bundles. C) Spindle shaped neoplastic cells forming interwoven bundles and the cells appeared as pleomorphic, hyperchromatic bizarre nuclei with abundant eosinophilic cytoplasm and 2-8 mitotic figures per high power filed (X400 H\u0026amp;E); (D) Spindle shaped neoplastic cells with pleomorphic, hyperchromatic bizarre nuclei with abundant eosinophilic cytoplasm forming interwoven bundles and infiltration of plasma cells, macrophages, eosinophils and tumor giant cells (X400 H\u0026amp;E).\u003c/p\u003e","description":"","filename":"2.png","url":"https://assets-eu.researchsquare.com/files/rs-4588055/v1/29a03fcf4f7f66770759d173.png"},{"id":59643719,"identity":"a5d2293b-ddc8-40e8-8300-6e31f252f3be","added_by":"auto","created_at":"2024-07-04 08:21:44","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":1441661,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-4588055/v1/bffb23d0-47cd-4e46-844b-682bc45ea086.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"Strobilocercus fasciolaris induced hepatic sarcoma in Wistar Rats","fulltext":[{"header":"1. Introduction","content":"\u003cp\u003eLaboratory animals, especially rodents such as rabbits, guinea pigs, mice, and rats, are widely utilized as models for investigating the pathobiology of various microbial or toxicological agents. The outcomes of these experimental studies can be significantly influenced by both biotic and abiotic factors, including environmental conditions, nutrition, and infection status [11, 27, 30]. Parasitic infections, in particular, can affect research results by altering the physiological and immunological characteristics of the hosts, potentially modifying their susceptibility to experimental stress. These alterations may lead to tissue damage, abnormal tissue growth, nutrient competition, blood volume reduction, and mechanical interference [15, 27].\u003c/p\u003e\n\u003cp\u003e\u0026nbsp;\u003cem\u003eTaenia taeniaeformis,\u0026nbsp;\u003c/em\u003ethough globally widespread, is rarely found in laboratory animals [29]. This parasite primarily infects wild and domestic carnivores, which serve as its definitive hosts. Rodents, such as rats, act as intermediate hosts, becoming infected by ingesting feed or water contaminated with the eggs from definitive hosts. Once ingested, the eggs hatch in the small intestine of the rodent, and the resulting embryos travel via the portal circulation to the liver, where they develop into the infective larval stage, known as \u003cem\u003eStrobilocercus fasciolaris\u003c/em\u003e or \u003cem\u003eCysticercus fasciolaris\u003c/em\u003e, or simply strobilocercus [32, 5]. Felines and other carnivorous definitive hosts become infected by consuming the liver of rats harboring the larval stage [29]. Another source of rodent infection is during the transportation of rats between the source and host institutions. Transportation stress often exacerbates these latent infections. As laboratory animal facilities often lack proper screening and quarantine measures for parasitic infestations, these infections can easily spread throughout the colony, infecting other animals\u003cem\u003e.\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003e\u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp; The pathology of strobilocercus-induced liver disease in rodents begins with the ingestion of \u003cem\u003eT. taeniaeformis\u003c/em\u003e eggs, which develop into larvae that form cysts in the liver parenchyma of rats, leading to progressive chronic inflammation and vigorous fibroplasia [6, 31]. Strobilocercus lodges in the liver parenchyma [6, 31], causing physiological and immunological changes, tissue damage, abnormal growth, nutrient competition, and deviations in animal health and research outcomes [15]. Additionally, fibroplastic and inflammatory lesions may progress to fibrosarcoma [20, 24]. There have been reports of the spontaneous occurrence of Taenid cyst-associated hepatic mesenchymal tumors with rare systemic metastasis and gastroenteropathy [10, 14, 20]. The parasite also poses a zoonotic risk to animal house workers, as humans can act as accidental hosts, with occasional reports of the parasite in the intestines and liver of infected individuals [12, 13, 36, 27].\u003c/p\u003e\n\u003cp\u003eThis study reports the incidental findings and pathology of Strobilocercus in laboratory rats, including the rare occurrence of associated hepatic sarcoma. Additionally, the study examines the potential impact on research outcomes, particularly in immunocompromised rats, and discusses its zoonotic implications.\u003c/p\u003e"},{"header":"2. Materials and methods","content":"\u003cp\u003e\u003cem\u003e2.1. Ethical Approval\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eThree-month-old Wistar rats were maintained in the university's laboratory animal house facility for another approved study (IAEC approval no. 110/IAEC/16). The animals were housed according to the guidelines for the care and use of animals in scientific research, as specified by the Institutional Animal Ethical Committee registered with the Committee for the Purpose of Control and Supervision of Experimental Animals (CPCSEA). During the study period, 23 rats died over the course of two weeks, displaying clinical signs of mild diarrhea, lethargy, and anorexia.\u003c/p\u003e\n\u003cp\u003e\u003cem\u003e2.2. Necropsy and sample collection \u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eA meticulous and scientific necropsy was performed on the deceased rats. All visceral organs were thoroughly inspected for the presence of cysts, adult parasites, or other gross lesions. For microscopic investigations, representative tissue samples containing parasitic cysts in situ, as well as some parasites, were carefully preserved in 10% neutral buffered formalin. All significant findings were documented with photographs.\u003c/p\u003e\n\u003cp\u003e\u003cem\u003e2.3. Morphological studies\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eThe liver samples, harboring parasitic cysts, were carefully isolated and submerged in a Petri dish filled with normal saline. The morphology of the developmental stages was then scrutinized for identification, involving the delicate dissection of the outer cyst covering from freshly collected livers [33]. A comprehensive gross morphological evaluation was carried out to ascertain the average number and size of the cysts. Subsequently, the cysts underwent precise dissection to assess the presence or absence of larval stages.\u003c/p\u003e\n\u003cp\u003e\u003cem\u003e2.4. Staining of developmental stages\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eThe liver lobes containing the cysts were washed in phosphate-buffered saline (1% PBS). The cyst wall was carefully dissected to collect the metacestode larva. The larva was then pressed between two glass slides and fixed in 10% neutral buffered formalin for 72-96 hours [7]. It was subsequently washed overnight in running tap water, stained with borax carmine, treated with acid alcohol, cleared in xylene, and finally mounted in dextrin plasticized xylene (DPX) (Meyer and Olsen, 1975). The development of scolices and the complete arrangement of the rostellar hooks and suckers were studied according to the method described by Meyer and Olsen (1975). Additionally, the larva was stained with H\u0026amp;E stain using the paraffin embedding technique for histopathology [23].\u003c/p\u003e\n\u003cp\u003e\u003cem\u003e2.5. Histopathology\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eRepresentative tissue samples from livers containing cysts, as well as intestines and individual parasites, were collected and fixed in 10% neutral buffered formalin. The developmental stages of the parasites and the tissues were then sectioned and processed for histopathological examination [23].\u003c/p\u003e"},{"header":"3. Results","content":"\u003cp\u003eTwenty-three deceased Wistar rats, aged 3 months, were submitted to the Department of Veterinary Pathology for necropsy and disease diagnosis. Grossly, cream-colored parasitic cysts were observed embedded in the hepatic parenchyma of 18 rats (78%) (Fig. 1A, 1B). On cut section, the metacestode larva was found inside the cysts (Fig. Inset 1B). Some rats had multiple parasitic cysts embedded in the liver parenchyma (Fig. 1B), containing parasites approximately 4 inches in length.\u003c/p\u003e\n\u003cp\u003e\u003cem\u003e3.1. Parasite and Larval Morphology\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eThe livers of the rats exhibited multiple cream-colored cysts affecting single or multiple lobes (Fig. 1A and 1B), ranging in size from 3-5mm. The recovered developmental stages measured between 4-20 inches in length, with relatively larger scolexes measuring 1-2 inches in length and a long neck (Fig. inset 1B). The larval body displayed pseudo segmentation throughout its length with a terminal bulged portion (Fig. Inset 1B). Stained parasites exhibited typical taenid morphology. In the larger cysts, well-developed scolices were observed (Fig. 1C), while most cysts were smaller in size. The larger scolices possessed two distinct rows of characteristic penknife-shaped hooks (Fig. 1C and 1D). A total of 42 hooks, consisting of 21 large and 21 small hooks, were recorded (Fig. Inset 1D). The mean size of the larger hooks was 390.89 ± 10.08 μm, and that of the smaller hooks was 241.46 ± 14.13 μm.\u003c/p\u003e\n\u003cp\u003e\u003cem\u003e3.2. Histopathology\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eMicroscopic examination of the affected liver sections revealed vacuolation, fatty changes, and inflammatory alterations with extensive infiltration of a mixed cellular population including lymphocytes, macrophages, and eosinophils (Fig. 2A and 2B). The hepatic tissue surrounding the cyst exhibited metaplastic changes and a proliferated fibrous connective tissue layer with strobilocercus within the cyst lumen (Fig. 2A and 2B). The cyst wall displayed a thin connective tissue capsule containing mature larger developmental stages and a thick wall of connective tissue in comparatively younger juvenile developmental stages. Furthermore, a zone of granulation tissue with lymphocytic cuffing was observed around certain cysts (Fig. 2A (black arrow) and 2B (black arrow)). Some liver sections showed hepatic parenchymal changes suggestive of neoplastic transformation of sarcoma (Fig. 2C and 2D). These neoplastic changes included poorly defined interwoven bundles of spindle cells that were pleomorphic, hyperchromatic, with bizarre nuclei, abundant eosinophilic cytoplasm, and separated by collagen as intercellular matrix (Fig. 2C and 2D). Infiltration of plasma cells, macrophages, and eosinophils surrounding the infected areas was also observed. Some sections suggestive of hepatic sarcomas exhibited tumor giant cells and 2-8 mitotic figures per high-power microscopic field (Fig. 2C (black arrow) and 2D (black arrow)). Hepatocytes in unaffected zones and at a distance from the cysts showed mild fatty changes and apparently healthy liver tissue. Hyperplasia of gastric mucosa, distension of gastric glands, and a marked increase in the number of goblet cells in the duodenal mucosa were also noted. No histopathological changes were observed in any other visceral organs.\u003c/p\u003e"},{"header":"4. Discussion","content":"\u003cp\u003e\u003cem\u003eTaenia taeniaeformis \u003c/em\u003eparasitizes the small intestine of both domestic and wild carnivores. Intermediate hosts, such as rodents, ingest the gravid proglottids passed in the feces of the definitive hosts. The parasite oncosphere traverses the stomach of the intermediate host and is released in their small intestine, penetrating the intestinal wall via the portal circulation and then carried to the liver, where it lodges into hepatic capillaries. Approximately six days post-infection, they are found inside small vesicles formed by the proliferation of host connective tissue on the liver surface. After a maturation period of 6-8 months, the developmental stages become fully grown infective strobilocerci with caudal vesicles. This parasite is cosmopolitan, with reports from various regions including India [20, 34], Japan [25], Egypt [35], and Mexico [29]. Experimental studies have revealed a relationship between the implantation of developmental stages and the development of liver tumors [8].\u003c/p\u003e\n\u003cp\u003eIn this study, Wistar rats exhibited signs of weight loss, lethargy, and mild diarrhea before succumbing. Similar clinical signs of gastroenteropathy have been documented previously [20, 16, 34]. Hepatic tumors induced \u003cem\u003eby T. taeniaeformis\u003c/em\u003e led to nonspecific clinical signs such as lethargy, weight loss, anorexia, and sudden death [14]. Infected rats displayed nonspecific clinicopathological changes that could significantly affect the outcomes of experimental studies. These changes may include elevated activity of liver-specific enzymes, decreased concentration of glucose, and an increase in neutrophils, lymphocytes, and eosinophils count in peripheral blood smears [4, 18].\u003c/p\u003e\n\u003cp\u003eOf the 23 rats submitted to the Department of Veterinary Pathology for necropsy, eighteen rats (18/23) were found to have cystic liver i.e., an incidence of 78%. The laboratory animal house maintains hygienic conditions and adheres to strict biosecurity measures; hence, the possible sources of infection could be the breeder colony or potential breaches in biosecurity protocols during transportation between the source and host institution.\u003c/p\u003e\n\u003cp\u003eAll rats were 3 months old, yet they exhibited cysts of varying sizes (3-5mm) and maturity levels. Gross morphological examination of the cysts revealed variability in number, size, and maturity, likely due to differences in exposure time and age at the time of infection. Another potential factor could be the variance in the immune response of individual rats to the parasite. Hanes [14] noted that rats with infections less than one month old typically had few immature cysts without developed scolices, affecting only 1-2 hepatic lobes. Conversely, rats with infections exceeding one month tended to have a higher number of cysts affecting the liver parenchyma more severely and involving a greater number of hepatic lobes.\u003c/p\u003e\n\u003cp\u003eThe scolex consistently contained two rows of characteristic penknife-shaped hooks with an equal number of large and small hooks in all cases. However, some researchers [2] have reported 30–40 hooks in the rostellum of the metacestode. Microscopic examination of affected liver sections revealed fatty, inflammatory, metaplastic changes in some areas, and proliferative fibrous tissue changes in the liver parenchyma. The infiltration of mononuclear cells in the hepatic parenchyma surrounding the cyst observed in this study aligns with previous reports [30, 20, 3, 22]. Histopathological findings were indicative of fibrosarcomatous changes characterized by spindle-shaped pleomorphic neoplastic cells in the liver parenchyma. Similar lesions have been reported in Sprague-Dawley rats [14, 20, 16]. Chronic infestation of larvae in the liver parenchyma induces chronic irritation and inflammatory changes, which can progress to metaplasia and ultimately neoplasia [1, 16]. The specific pathogenesis of neoplastic changes in the liver is not fully understood, but a causal association of \u003cem\u003eT. taeniaeformis\u003c/em\u003e with hepatic tumors is well established [20, 16]. The causal hypothesis suggests the involvement of the parasite, larval-derived oncogenic substances, chronic inflammation, irritation of liver parenchyma due to the presence of larval cysts, oxidative damage, and immunosuppression caused by the adult parasite and larvae themselves [8, 9, 14]. Histopathological changes in the gastrointestinal tract (GIT) include hyperplasia of mucosal glands and hypertrophy of the mucosa. Infected rats may develop lesions of inhibited gastric acid secretion, hypergastrinemia, and gastric and intestinal mucosal hyperplasia [18, 26]. These changes are attributed to larval secretions in the gastric and intestinal lumen [21].\u003c/p\u003e"},{"header":"5. Conclusion ","content":"\u003cp\u003eThe current study documents \u003cem\u003eStrobilocercus fasciolaris\u003c/em\u003e infection in Wistar rats, leading to the subsequent development of hepatic sarcoma. The developmental stage of the parasite acts as a foreign body, causing gastroenteropathy and chronic irritation and inflammation of the liver parenchyma, ultimately resulting in neoplasia. This report underscores the importance of screening laboratory animals for parasitic infestations and implementing quarantine measures before introducing them into the colony. Additionally, this infection presents a small but significant zoonotic risk to laboratory animal house workers and animal handlers. Infection can occur through contamination of clothing and other personal items while handling infected animals, their feed, feeders, utensils, bedding materials, etc., or due to breaches in biosecurity protocols.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eAcknowledgments\u003c/strong\u003e\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eAuthors are thankful to Head, Department of Veterinary Microbiology, Department of Veterinary Pathology and Dean, College of Veterinary Science and Animal Husbandry for providing the facilities to conduct this study.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthor\u0026rsquo;s contribution\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eRK performed the histopathological examination and wrote the manuscript.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eVK identified the parasite and edited the final draft.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eSource of funding\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eData accessibility statement\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEthical statement\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u0026nbsp;The study was ethically approved by Institutional Animal Ethical Committee (IAEC) approval no. 110/IAEC/16.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConsent for publication\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConflict of interest\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNone of the authors is having conflict of interest.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eAl-Jashamy, K., El Salihi, K., Sheikh, A. and Saied, H., 2004, May. 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Public health concerns of Taenidae and their metacestodes. \u003cem\u003eTrop Med Surg\u003c/em\u003e, \u003cem\u003e3\u003c/em\u003e(1), p.e123.\u003c/li\u003e\n\u003cli\u003ePerec-Matysiak, Agnieszka, Anna Okulewicz, Joanna Hildebrand, and Grzegorz Zaleśny. 2006. Helminth parasites of laboratory mice and rats. \u003cem\u003eWiadomości Parazytologiczne,\u003c/em\u003e 52 (2), 99-102.\u003c/li\u003e\n\u003cli\u003eRodr\u0026iacute;guez-Vivas, R.I., Panti-May, J.A., Parada-L\u0026oacute;pez, J., Hern\u0026aacute;ndez-Betancourt, S.F. and Ruiz-Pi\u0026ntilde;a, H.A., 2011. The occurrence of the larval cestode Cysticercus fasciolaris in rodent populations from the Cuxtal ecological reserve, Yucatan, Mexico. \u003cem\u003eJournal of Helminthology\u003c/em\u003e, \u003cem\u003e85\u003c/em\u003e(4), pp.458-461. \u003c/li\u003e\n\u003cli\u003eSingla, Lachhman D., Neena Singla, Vir R. Parshad, Prayag D. Juyal, and Naresh K. Sood. 2008. Rodents as reservoirs of parasites in India.\u0026quot; \u003cem\u003eIntegrative zoology,\u003c/em\u003e 3 (1), 21-26. \u003c/li\u003e\n\u003cli\u003eSingh, Vinod Kumar, Rahul Kumar, Amit Kumar, and Sharad Kumar Yadav., 2008. Microbiology and Histo-Pathology of Mandibulofacial Abscess in A BALB/c Mouse. \u003cem\u003eApplied Biological Research,\u003c/em\u003e 20 (2), 218-220.\u003c/li\u003e\n\u003cli\u003eSinniah, Bharathalingam, Muniandy Narasiman, Saequa Habib, and Ong Gaik Bei., 2014. Prevalence of Calodium hepaticum and Cysticercus fasciolaris in urban rats and their histopathological reaction in the livers. \u003cem\u003eJournal of Veterinary Medicine\u003c/em\u003e, Article-ID. 172829\u003c/li\u003e\n\u003cli\u003eSoulsby, Ernest Jackson Lawson., 1982. \u003cem\u003eHelminths, arthropods and protozoa of domesticated animals\u003c/em\u003e. No. 7th edition. Bailliere Tindall, 10 Greycoat Place.\u003c/li\u003e\n\u003cli\u003eThangapandiyan, M., Balachandran, C., Preetha, S.P., Mohanapriya, T., Nivethitha, R., Pavithra, S. and Sridhar, R., 2017. Gross, histopathological and immunohistochemical study on strobilocercus of Taenia taeniaeformis infection in the liver of laboratory rats (Rattus norvegicus) in India. Veterinary Parasitology: Regional Studies and Reports, 10, pp.35-38. \u003c/li\u003e\n\u003cli\u003eYounis, A.E., Saad, A.I., El-Akhal, I.R.M. and Saleh, N.M.K., 2021. A parasitological survey of zoonotic cestodes carried by house rats in Aswan, Egypt, reveals cryptic diversity at the molecular level. \u003cem\u003eVeterinary World\u003c/em\u003e, \u003cem\u003e14\u003c/em\u003e(8), p.2160. \u003c/li\u003e\n\u003cli\u003eZibaei, Mohammad, Seyed Mahmoud Sadjjadi, and Bahador Sarkari., 2007. Prevalence of Toxocara cati and other intestinal helminths in stray cats in Shiraz, Iran. \u003cem\u003eTrop Biomed,\u003c/em\u003e 24 (2), 39-43.\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"Hepatic sarcoma, Strobilocercus fasciolaris, Wistar rats, Histopathology","lastPublishedDoi":"10.21203/rs.3.rs-4588055/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-4588055/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003e\u003cstrong\u003eBackground \u003c/strong\u003e\u003cem\u003eTaenia taeniaeformis \u003c/em\u003einfects the small intestine of domestic and wild carnivores. Rodents, the intermediate hosts, ingest the gravid proglottids from the carnivores' feces. The parasite’s oncosphere reaches the liver via portal circulation after penetrating the wall of small intestine.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003ePurpose\u003c/strong\u003e The present study reports hepatic sarcoma caused by strobilocercus- the metacestodal stage of \u003cem\u003eTaenia taeniformis\u003c/em\u003e in laboratory rats.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMethod\u003c/strong\u003e Twenty-three rats in a colony of 72 male wistar rats died suddenly with clinical signs of weakness, lethargy and mild diarrhea. The carcasses were sent to the department of veterinary pathology for postmortem examination.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eResults\u003c/strong\u003e Grossly, the liver of 18 rats were having multiple, cream to white cysts of variable sizes of \u003cem\u003eStrobilocercus fasciolaris\u003c/em\u003e. Histopathologically, the hepatic tissue surrounding the cysts revealed zones of fatty change, inflammation with infiltration of plasma cells, macrophages and eosinophils, presence of granulation tissue, metaplasia and fibrosarcoma. The pathological changes in the stomach and small intestines include hyperplasia of gastric and intestinal glands and their mucosa.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConclusion\u003c/strong\u003e The likelihood of altered research results of studies conducted in immune-compromised \u003cem\u003eT. taeniformis\u003c/em\u003einfected rats and its zoonotic implications are discussed herewith.\u003c/p\u003e","manuscriptTitle":"Strobilocercus fasciolaris induced hepatic sarcoma in Wistar Rats","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2024-07-03 05:21:09","doi":"10.21203/rs.3.rs-4588055/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"8bad9bac-b2db-41a2-83d8-5ee897087186","owner":[],"postedDate":"July 3rd, 2024","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2024-07-04T08:21:32+00:00","versionOfRecord":[],"versionCreatedAt":"2024-07-03 05:21:09","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-4588055","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-4588055","identity":"rs-4588055","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}