Ultrastructural aspects of the host cellular immune response to Taenia crassiceps metacestodes.

When three Taenia crassiceps metacestodes were injected intraperitoneally into C3H mice primed by previous subcutaneous inoculation of metacestodes, larvae which were resistant to early immune damage by the humoral response were encapsulated by host cells and rejected. Initially, normal larvae were encapsulated primarily by eosinophils and macrophages. In the early stages of encapsulation, both cell types showed severe degenerative changes and disruption of cell membranes, but there was no evidence of tegumental damage to the encapsulated larvae. Later, mast cells appeared in the capsules surrounding the larvae. After mast cells became common, all of the cell types present were normal, and damage to the larval Tegument became apparent. Ultimately, interaction of eosinophils, mast cells, macrophages, and lymphocytes resulted in death of the encapsulated larvae. These results suggest that larvae may secrete substances toxic to host cells, and that mast cells are necessary for rejection of larvae.     

Taenia crassiceps: effect of normal and immune serum on metacestodes in vitro.

The effect of normal and immune serum on Taenia crassiceps larvae in vitro was assessed by Evans blue dye uptake and electron microscopy. Normal guinea pig, rabbit, goat, and fetal calf serum did not have any significant detrimental effects upon the larvae after 7 days of culture in vitro. Culture for 7 days in normal mouse serum resulted in some loss of tegumental microtriches but the tegument itself remained intact. Culture in hyperimmune rabbit serum resulted in complete loss of the tegument and disruption of subtegumental structures within 48 hr. The effects of immune mouse serum in vitro closely paralleled those previously seen during early immune damage in vivo. Immune serum taken 2 to 4 weeks after secondary intraperitoneal infection with T. crassiceps metacestodes caused loss of the larval tegument and degeneration of the subtegumental tissues after 7 days in culture, whereas immune mouse serum taken 6 weeks after secondary infection caused only minor ultrastructural changes and appeared to be less toxic to larvae than normal mouse serum. Although complement appeared to increase the number and severity of the tegumental lesions, the presence of heat-labile components of complement was not essential for mediation of tegumental damage by immune mouse serum.     

Kinetics of primary and secondary infections with Taenia crassiceps metacestodes (Zeder, 1800) rudolphi, 1810 (cestoda: cyclophyllidea)

Siebert A. E. Jr., Good A. H. & Simmons J. E. 1978. Kinetics of primary and secondary infections with Taenia crassiceps metacestodes (Zeder, 1800) Rudolphi, 1810 (Cestoda: Cyclophyllidea). International journal for Parasitology8: 39–43. When three T. crassiceps metacestodes were inoculated intraperitoneally in mice as a primary infection, approximately 50% of the larvae recovered during the first 4 weeks after inoculation were found to be dead, while in mice primed by previous subcutaneous inoculation, about 85% of the larvae died. Larvae which survived the first 4 weeks following primary intraperitoneal inoculation reproduced asexually by exogenous budding and produced viable infections within the host mice. But larvae in secondary infections were encapsulated by host granulomata, failed to reproduce asexually, and did not produce viable infections. In mice given intraperitoneal inoculations of seven, ten and twenty metacestodes, fewer larvae were killed and little encapsulation response was noted, though host cells were common at the budding region of the larvae. Such a biphasic host-response to the infection has not previously been reported for larval cestode infections, and the reduction in host response associated with increased worm burdens may indicate possible depression of the host immune system.     

Taenia crassiceps: Immunity to metacestodes in BALB/c and BDF1 mice

The kinetics of primary and secondary infections with Taenia crassiceps larvae and the effects of immune serum on T. crassiceps larvae were studied in BALB/c and BDF1 mice. In both strains of mice a substantial degree of resistance to reinfection comparable to that previously reported in C3H mice can be induced by subcutaneous injection of three larvae 3 weeks prior to intraperitoneal challenge infection. Both early immune damage in the absence of adherent host cells and encapsulation by host cells are involved in rejection of larvae by BALB/c and BDF1 mice, but in both of these strains early immune damage is less pronounced and the cellular encapsulation response considerably more prominent than in the C3H mice studied previously. This difference is also reflected in the effect of immune serum on T. crassiceps metacestodes in vitro: immune serum from BALB/c and BDF1 mice is less effective than immune serum taken from C3H mice at comparable times after challenge infection in mediating damage to T. crassiceps larvae in vitro in the absence of host cells. These results suggest that genetically determined differences in immune capability can alter the state of equilibrium existing among different immune effector mechanisms without producing measurable effects upon overall host resistance to reinfection.     

Cross-protection between the metacestodes of Mesocestoides corti and Taenia crassiceps in mice

Novak M. 1984. Cross-protection between the metacestodes of Mesocestoides corti and Taenia crassiceps in mice. International Journal for Parasitology14: 497–501. Infection with M. corti generated significant resistance against a challenge with T. crassiceps introduced either 2 or 6 weeks after primary infection. Challenge infection with T. crassiceps did not influence primary infection with M. corti. Infection with T. crassiceps protected significantly against challenge with M. corti given 2 weeks but not 6 weeks after the primary infection. Challenge infection with M. corti significantly suppressed primary infection with T. crassiceps.     

A single oral treatment with mebendazole for the control of Taenia crassiceps larval infections in rats.

A single oral treatment with mebendazole for the control of Taenia crassiceps larval infections in rats. International Journal for Parasitology9: 73–76. Rats infected with Taenia crassiceps larvae were treated with mebendazole. At a sublethal dose level of 50 mg/kg, a single large oral treatment proved to be markedly more effective in killing cysts than the same amount of drug divided into 10 daily smaller doses. Levamisole promoted a vigorous host cellular response to the intraperitoneal cysts, but when incorporated with mebendazole, it did not enhance the action of the latter drug.     

Taenia larvae possess distinct acetylcholinesterase profiles with implications for host cholinergic signalling

Larvae of the cestodes Taenia solium and Taenia crassiceps infect the central nervous system of humans. Taenia solium larvae in the brain cause neurocysticercosis, the leading cause of adult-acquired epilepsy worldwide. Relatively little is understood about how cestode-derived products modulate host neural and immune signalling. Acetylcholinesterases, a class of enzyme that breaks down acetylcholine, are produced by a host of parasitic worms to aid their survival in the host. Acetylcholine is an important signalling molecule in both the human nervous and immune systems, with powerful modulatory effects on the excitability of cortical networks. Therefore, it is important to establish whether cestode derived acetylcholinesterases may alter host neuronal cholinergic signalling. Here we make use of multiple techniques to profile acetylcholinesterase activity in different extracts of both Taenia crassiceps and Taenia solium larvae. We find that the larvae of both species contain substantial acetylcholinesterase activity. However, acetylcholinesterase activity is lower in Taenia solium as compared to Taenia crassiceps larvae. Further, whilst we observed acetylcholinesterase activity in all fractions of Taenia crassiceps larvae, including on the membrane surface and in the excreted/secreted extracts, we could not identify acetylcholinesterases on the membrane surface or in the excreted/secreted extracts of Taenia solium larvae. Bioinformatic analysis revealed conservation of the functional protein domains in the Taenia solium acetylcholinesterases, when compared to the homologous human sequence. Finally, using whole-cell patch clamp recordings in rat hippocampal brain slice cultures, we demonstrate that Taenia larval derived acetylcholinesterases can break down acetylcholine at a concentration which induces changes in neuronal signalling. Together, these findings highlight the possibility that Taenia larval acetylcholinesterases can interfere with cholinergic signalling in the host, potentially contributing to pathogenesis in neurocysticercosis.     

Echinococcus multilocularis in the mouse: The in vitro protoscolicidal activity of peritoneal macrophages

Baron R. W. and Tanner C. E. 1977. Echinococcus multilocularis in the mouse: the in vitro protoscolicidal activity of peritoneal macrophages. International Journal for Parasitology7: 489–495. The larvae of Echinococcus multilocularis are susceptible to the protoscolicidal activity of infected A/J mouse peritoneal cells. It is shown that the effector cell in this response is an activated macrophage. Preincubation of protoscolices in ‘immune’ serum increases their susceptibility to the protoscolicidal activity of infected mouse peritoneal cells. Macrophages activated nonspecifically by BCG or Taenia crassiceps infections also exhibit protoscolicidal activity in vitro. The identity of the effector cell was confirmed by scanning electron microscopy. It is shown that ‘immune’ macrophages adhere to and form close cellular contacts with the protoscolex surface. It is concluded that resistance to hydatid infections is mediated by activated macrophages.     

Taenia crassiceps in the rat: differences in susceptibility to infection and development of immunocompetence in relation to age and host strain.

The influence of host strain and age upon susceptibility of rats to infection with Taenia crassiceps metacestodes was examined. The polyclonal T-cell activators concanavalin-A and phytohaemagglutinin were used to demonstrate that the onset of host resistance to infection with the cestode is coincident with the acquisition of responsiveness to mitogens in the post-natal maturation of the AS₂ rat.     

Peripheral Taenia infection increases immunoglobulins in the central nervous system

Human cysticercosis is a disease caused by larvae of the cestode Taenia solium. It is an important common cause of adult-onset seizures world-wide where it exacts a debilitating toll on the health and well-being of affected communities. It is commonly assumed that the major symptoms associated with cysticercosis are a result of the direct presence of larvae in the brain. As a result, the possible effects of peripherally located larvae on the central nervous system are not well understood. To address this question, we utilised the Taenia crassiceps intra-peritoneal murine model of cysticercosis, where larvae are restricted to the peritoneal cavity. In this model, previous research has observed behavioural changes in rodents but not the development of seizures. Here we used ELISAs, immunoblotting and the Evans Blue test for blood–brain barrier permeability to explore the central effects of peripheral infection of mice with T. crassiceps. We identified high levels of parasite-targeting immunoglobulins in the sera of T. crassiceps-infected mice. We show that the T. crassciceps larvae themselves also contain and release host immunoglobulins over time. Additionally, we describe, for the first known time, significantly increased levels of IgG within the hippocampi of infected mice, which are accompanied by changes in blood–brain barrier permeability. However, these T. crassiceps-induced changes were not accompanied by alterations to the levels of proinflammatory, pro-seizure cytokines in the hippocampus. These findings contribute to the understanding of systemic and neuroimmune responses in the T. crassiceps model of cysticercosis, with implications for the pathogenesis of human cysticercosis.     

Subcutaneous cysticercosis due to Taenia crassiceps (Cestoda: Taeniidae) in an imported steppe lemming in Japan

This study describes a subcutaneous proliferative cysticercosis in a pet steppe lemming, Lagurus lagurus (Rodentia: Cricetidae), bred and imported from Czech Republic into Japan. Numerous metacestodes were collected from the subcutaneous cystic lesion of the left medial thigh. Four surgical removals were coupled with anthelmintic treatment but ended with recurrence. Based on morphological features and mitochondrial DNA sequences, the metacestodes were identified as the larval stage of Taenia crassiceps (Zeder, 1800). This is the first case of infection with larval T. crassiceps in rodents of the genus Lagurus, and becomes the third case of the parasite detected from imported animals in Japan. Related public health concerns are discussed.     

Taenia crassiceps: Regional variations in ultrastructure and evidence of endocytosis in the cysticercus' tegument

Regional variations in the thickness of the tegument, in the morphology of microtriches and mitochondria, in the distribution of dense bodies, smooth micropinocytotic vesicles (SMVs), and coated micropinocytotic vesicles (CMVs) have been shown for the cysticercus of Taenia crassiceps. The number of SMVs and CMVs present in the syncytial layer are in inverse proportion to each other, the former being more numerous in the bladder wall and upper part of the invagination canal and the latter in the lower part of the canal and the rostellar region. Tegumental cells contain numerous granular endoplasmic reticulum-Golgi complexes involved in the synthesis of both primary lysosomes and dense bodies. Vesicles characteristic of various stages of heterolysosomes are present and show regional variations in numbers and size. Acid phosphatase activity (EC 3.1.3.2) is present on the tegumental surface, and within the Golgi complex, primary lysosomes, and heterophagosomes of the tegumental cells. CMVs are reported for the first time in the tegument of any helminth and have characteristics similar to CMVs in other tissues. T. crassiceps, therefore, because of the presence of both SMVs and CMVs, is a unique model system for the study of basic mechanisms of endocytosis.     

Characterization of S3Pvac Anti-Cysticercosis Vaccine Components: Implications for the Development of an Anti-Cestodiasis Vaccine

Background

Cysticercosis and hydatidosis seriously affect human health and are responsible for considerable economic loss in animal husbandry in non-developed and developed countries. S3Pvac and EG95 are the only field trial-tested vaccine candidates against cysticercosis and hydatidosis, respectively. S3Pvac is composed of three peptides (KETc1, GK1 and KETc12), originally identified in a Taenia crassiceps cDNA library. S3Pvac synthetically and recombinantly expressed is effective against experimentally and naturally acquired cysticercosis.

Methodology/Principal Findings

In this study, the homologous sequences of two of the S3Pvac peptides, GK1 and KETc1, were identified and further characterized in Taenia crassiceps WFU, Taenia solium, Taenia saginata, Echinococcus granulosus and Echinococcus multilocularis. Comparisons of the nucleotide and amino acid sequences coding for KETc1 and GK1 revealed significant homologies in these species. The predicted secondary structure of GK1 is almost identical between the species, while some differences were observed in the C terminal region of KETc1 according to 3D modeling. A KETc1 variant with a deletion of three C-terminal amino acids protected to the same extent against experimental murine cysticercosis as the entire peptide. On the contrary, immunization with the truncated GK1 failed to induce protection. Immunolocalization studies revealed the non stage-specificity of the two S3Pvac epitopes and their persistence in the larval tegument of all species and in Taenia adult tapeworms.

Conclusions/Significance

These results indicate that GK1 and KETc1 may be considered candidates to be included in the formulation of a multivalent and multistage vaccine against these cestodiases because of their enhancing effects on other available vaccine candidates.     

Interactions between Taenia taeniaeformis and host cells in vitro: rapid adherence of peritoneal cells to strobilocerci

Engelkirk P. G., Williams J. F. and Signs M. M. 1981. Interactions between Taenia taeniaeformis and host cells in vitro: rapid adherence of peritoneal cells to strobilocerci. International Journal for Parasitology11: 463–474. Strobilocerci of Taenia taeniaeformis, incubated for l h in vitro with various combinations of serum and peritoneal cells from infected or non-infected rats, were examined at the ultrastructural level for evidence of cell adherence and tegumental damage. Maximal adherence and surface alterations occurred when larvae were incubated in the presence of cells and fresh serum. This was true regardless of whether the cells or the serum had been obtained from infected or non-infected donors. No tegumental damage was seen when parasites were incubated with or without cells in the absence of serum. Serum enhancement was either much reduced or abolished by heat treatment (56°C for 1 h). In the presence of EDTA, tegumental lesions still developed, but adherence of cells, especially those from non-infected rats, decreased markedly. The predominant cells interacting with the larval surface were eosinophils; these took up parasite material within phagosomes and appeared to strip microtriches from the tegumental free surface. Mast cells, some of which became degranulated, were also present in the adherent cell masses. The results indicate that potent non-specific effector mechanisms can rapidly damage the tegument of T. taeniaeformis, in vitro, in contrast to the failure of recognition and rejection by host defenses in vivo. Established strobilocerci are therefore not invulnerable but the balance of the host-parasite relationship in vivo must favor their survival.     

Oestradiol and progesterone differentially alter cytoskeletal protein expression and flame cell morphology in Taenia crassiceps

We examined the effects of oestradiol (E₂) and progesterone (P₄) on cytoskeletal protein expression in the helminth Taenia crassiceps — specifically actin, tubulin and myosin. These proteins assemble into flame cells, which constitute the parasite excretory system. Total protein extracts were obtained from E₂- and P₄-treated T. crassiceps cysticerci and untreated controls, and analysed by one- and two-dimensional protein electrophoresis, flow cytometry, immunofluorescence and videomicroscopy. Exposure of T. crassiceps cysticerci to E₂ and P₄ induced differential protein expression patterns compared with untreated controls. Changes in actin, tubulin and myosin expression were confirmed by flow cytometry of parasite cells and immunofluorescence. In addition, parasite morphology was altered in response to E₂ and P₄ versus controls. Flame cells were primarily affected at the level of the ciliary tuft, in association with the changes in actin, tubulin and myosin. We conclude that oestradiol and progesterone act directly on T. crassiceps cysticerci, altering actin, tubulin and myosin expression and thus affecting the assembly and function of flame cells. Our results increase our understanding of several aspects of the molecular crosstalk between host and parasite, which might be useful in designing anthelmintic drugs that exclusively impair parasitic proteins which mediate cell signaling and pathogenic reproduction and establishment.     

Damage and early destruction of Taenia taeniaeformis larvae in resistant hosts, and anomalous development in susceptible hosts: a light microscopic and ultrastructural study

Bortoletti G., Conchedda M. and Ferretti G. 1985. Damage and early destruction of Taenia taeniaeformis larvae in resistant hosts, and anomalous development in susceptible hosts: a light microscopic and ultrastructural study. International Journal for Parasitology15: 377–384. Taenia taeniaeformis larvae in resistant C57 mice have been studied from 5th to 15th day post-infection (L5–L15) both at the light and electron microscopic level. L5 stages were already damaged and total destruction occurred by approx. 15 days post-infection. In stage L5, unlike fertile larvae from C3H mice, the perilarval amorphous layer (PAL) was generally absent, and the host's cells were in close contact with the parasite surface. At this stage eosinophils were already present together with neutrophils and macrophages. Larvae were seen increasing in volume between stages L6 and L8, but remained constant from stages L9 to L14, while both the tegumental distal cytoplasm (TDC) and the subtegumental cellular layer (SCL) gradually decreased. In stages L10–L14 only a narrow TDC separated the larval cavity from host cells. After the larval tegument had been reduced in thickness the eosinophil lytic enzyme release onto the parasite surface contributed to produce a ‘hole’ in the TDC where host cells penetrated and gradually filled the larval cavity of L15, destroying the parasitic residues. Therefore anomalous small larvae (L50 and more) from C3H mice (susceptible host) have been studied: in these the scolex anlagen was absent or greatly reduced; the TDC was very narrow and the SCL greatly damaged. Outside the larva the ‘host tissue’ appeared as an unidentifiable amorphous material. These larvae cannot be considered ‘dead’ but are defined as sterile.     

Androgens Exert a Cysticidal Effect upon Taenia crassiceps by Disrupting Flame Cell Morphology and Function

The effects of testosterone (T₄) and dihydrotestosterone (DHT) on the survival of the helminth cestode parasite Taenia crassiceps, as well as their effects on actin, tubulin and myosin expression and their assembly into the excretory system of flame cells are described in this paper. In vitro evaluations on parasite viability, flow cytometry, confocal microscopy, video-microscopy of live flame cells, and docking experiments of androgens interacting with actin, tubulin, and myosin were conducted. Our results show that T₄ and DHT reduce T. crassiceps viability in a dose- and time-dependent fashion, reaching 90% of mortality at the highest dose used (40 ng/ml) and time exposed (10 days) in culture. Androgen treatment does not induce differences in the specific expression pattern of actin, tubulin, and myosin isoforms as compared with control parasites. Confocal microscopy demonstrated a strong disruption of the parasite tegument, with reduced assembly, shape, and motion of flame cells. Docking experiments show that androgens are capable of affecting parasite survival and flame cell morphology by directly interacting with actin, tubulin and myosin without altering their protein expression pattern. We show that both T₄ and DHT are able to bind actin, tubulin, and myosin affecting their assembly and causing parasite intoxication due to impairment of flame cell function. Live flame cell video microscopy showing a reduced motion as well changes in the shape of flame cells are also shown. In summary, T₄ and DHT directly act on T. crassiceps cysticerci through altering parasite survival as well as the assembly and function of flame cells.     

Identification and culture of proliferative cells in abnormal Taenia solium larvae: Role in the development of racemose neurocysticercosis

Racemose neurocysticercosis is an aggressive disease caused by the aberrant expansion of the cyst form of Taenia solium within the subarachnoid spaces of the human brain and spinal cord resulting in a mass effect and chronic inflammation. Although expansion is likely caused by the proliferation and growth of the parasite bladder wall, there is little direct evidence of the mechanisms that underlie these processes. Since the development and growth of cysts in related cestodes involves totipotential germinative cells, we hypothesized that the expansive growth of the racemose larvae is organized and maintained by germinative cells. Here, we identified proliferative cells expressing the serine/threonine-protein kinase plk1 by in situ hybridization. Proliferative cells were present within the bladder wall of racemose form and absent from the homologous tissue surrounding the vesicular form. Cyst proliferation in the related model species Taenia crassiceps (ORF strain) occurs normally by budding from the cyst bladder wall and proliferative cells were concentrated within the growth buds. Cells isolated from bladder wall of racemose larvae were established in primary cell culture and insulin stimulated their proliferation in a dose-dependent manner. These findings indicate that the growth of racemose larvae is likely due to abnormal cell proliferation. The different distribution of proliferative cells in the racemose larvae and their sensitivity to insulin may reflect significant changes at the cellular and molecular levels involved in their tumor-like growth. Parasite cell cultures offer a powerful tool to characterize the nature and formation of the racemose form, understand the developmental biology of T. solium, and to identify new effective drugs for treatment.     

Morphological studies on the early development of Taenia taeniaeformis larvae in susceptible mice

Bortoletti G. and Ferretti G. 1985. Morphological studies on the early development of Taenia taeniaeformis larvae in susceptible mice. International Journal for Parasitology15: 365–375. Taenia taeniaeformis larvae which develop into infective strobilocerci in C3H mice have been studied from the 5th to the 15th day of development (L5–L15), both at light and electron microscope level. The L5 were initially compact, without a central cavity but then become vacuolized. Until stages L7–L8 they were surrounded by a perilarval amorphous layer (PAL) made up of a finely granular material which prevented the host cells from making contact with the larval tegument. The larval volume increased considerably between stages L6 and L8, remained unchanged from L9 to L13, but continued to increase thereafter. The larval cellular layer, which appeared as a single, large ‘syncitial system’, grow until stages L14–L15 when the scolex anlagen began to form. The tegument was initially incompletely organized and was covered by microvilli. These were completely replaced by microtriches from stage L8 onward. Sometimes both microvilli and microtriches were together observed in stage L7. Microvilli fragments, sometimes beaded, could be observed at L5 within the damaged cytoplasm of host cell debris. Very often they were branched at different heights, especially in stages L5–L7. In L10–L15 all undamaged microtriches increased in density and formed bundles which invaded the host cells. In stages L5–L8 and in some L9, muscular bundles started to become organized inside the tegumental distal cytoplasm (TDC), and after become independent in the subtegumental cellular layer (SCL). Until L8–L9 the larvae were surrounded by host cells debris. From stages L8–L10 onwards the adjacent host cells were less damaged though the larval microtriches penetrated them deeply. In stages L5–L7 neutrophils together with macrophages and some damaged hepatocytes were detected, while eosinophils were present only from L8 onward. In the other stages neutrophils clearly diminished in numbers, whereas macrophages had increased. No mastcells and few plasma cells were observed.     

Steroid hormone production by parasites: the case of Taenia crassiceps and Taenia solium cysticerci

Many examples of reciprocal endocrine interactions between parasites and hosts have been found in insects, arthropods and mammals. Cysticercosis produced by Taenia solium metacestodes is a widely distributed parasite infection that affects the human and the pig. Taenia crassiceps experimental murine cysticercosis has been used to explore the role of biological factors involved in host–parasite interactions. We had shown that T. crassiceps cysticercosis affects the serum concentration of steroid hormones and the reproduction behavior of the male mice host. In an effort to understand the biology of the parasite, we had investigated the parasite capacity to produce sex steroids. For this purpose, T. crassiceps cysticerci were incubated in the presence of different steroid precursors. TLC and recrystallization procedures showed that testosterone is produced from ³H-androstenedione in cysticerci. The conversion of ³H-testosterone to androstenedione, although present is much less significant. In addition, we had studied the production of testosterone by T. solium cysticerci. For this purpose, cysticerci were dissected from pork meat and incubated as above described. The results showed that T. solium cysticerci also produce testosterone. We have speculated about the importance of androgens in the growth of T. crassiceps cysticerci and found that the addition of the antiandrogen flutamide to the culture media of the parasites significantly decreased ³H-thymidine incorporation. We therefore hypothesized, that the ability of cysticerci to produce testosterone from steroid precursors might be important for the parasite growth and development.