Suppression of host Th1-type granulomatous inflammation by Taenia solium metacestodes is related to down-regulation of osteopontin gene expression

Inflammation and granuloma formation in human neurocysticercosis has been attributed to Th1-type immune responses of the host. In the present murine model, over 94% of Taenia solium metacestodes were viable and elicited no granulomatous inflammation, whereas parasites killed by praziquantel treatment elicited rapid granuloma formation that calcified within 2weeks. Osteopontin (OPN) is a Th1-related cytokine that is up-stream of IL-12 and which may play an essential role in granuloma formation and calcification. OPN mRNA expression was down-regulated in tissues surrounding viable cysticerci, but was up-regulated in inflammatory tissues surrounding degenerating cysticerci. Moreover, co-culture with a viable cysticercus or ES products from these metacestodes led to a decrease in OPN, IFN-gamma and IL-12 expression, whereas co-culture with somatic proteins enhanced OPN expression by leukocytes. Addition of recombinant mouse OPN (rmOPN) counteracted the down-regulation of IL-12 and IFN-gamma mRNA expression, but not OPN mRNA expression, in leukocyte cultures. Furthermore, injection of rmOPN into the tissues surrounding implanted cysticerci enhanced inflammatory responses while a similar injection of an anti-rmOPN antibody reduced inflammation. These findings suggest that the suppression of host Th1-type granulomatous inflammation by ES products from T. solium metacestodes is related to down-regulation of OPN gene expression.     

Oncospheres of Taenia solium and T. saginata asiatica develop into metacestodes in normal and immunosuppressed mice.

Normal and immunosuppressed mice were infected with oncospheres of Taenia saginata asiatica and T. solium. Although normal ICR mice were not susceptible to these two parasites, cysticerci were recovered from the immunosuppressed ones following venous injection. For T. s. asiatica, immunosuppressed ICR mice had an infection rate of 12.5% and six cysticerci of this parasite were recovered from three males. After injection of T. solium oncospheres, a high infection rate of 57% was obtained and 23 cysticerci were collected from 13 male immunosuppressed ICR mice. The immunosuppressed C57 mice had the highest infection rate (100%) and cysticercus recovery rate (2.4%) for T. solium. The infection rate and cysticercus recovery rate in six normal C57 mice were 40% and 3% respectively. The immunosuppressed ICR, Balb/c and C3H mice were also susceptible to T. s. asiatica.     

Detection of genetic variation in Taenia solium

Genetic variability among Taenia solium isolates was studied in 160 cysticerci from 6 pigs, 4 from Mexico, 1 from Honduras, and 1 from Tanzania. Random amplified polymorphic DNA (RAPD) analysis performed with 4 commercial primers showed 88% polymorphic loci and an average heterozygosity of 0.077; however, several alleles were fixed within each isolate. Linkage disequilibrium analysis indicated that 3 of the 6 isolates had a random association of alleles, whereas the other 3 had a clonal structure. These results suggest the existence of local lineages in T. solium, with events of genetic recombination within them.     

Vaccination against Taenia solium cysticercosis in pigs using native and recombinant oncosphere antigens.

Pigs were immunised with antigens derived from Taenia solium oncospheres or with a pool of three recombinant antigens from Taenia ovis, and subsequently challenged with T. solium eggs. The native oncosphere antigens induced 83% protection against viable, and 89% protection against the total number of cysticerci established following the challenge infection. Immunisation with the recombinant T. ovis antigens induced 93% protection against the establishment of viable cysticerci, and 74% protection against the total number of cysticerci. These results, and those achieved elsewhere with Taenia saginata and T. ovis, support the possibility of developing a practical vaccine to assist in the control of transmission of T. solium through pigs.     

In situ detection of antigenic glycoproteins in Taenia solium metacestodes

Taenia solium has a complex life cycle. Its cysticercus can lodge in the brain, causing neurocysticercosis (NCC), and the adult tapeworm's survival in the intestine results in taeniasis. In this study, the in situ detection of previously described glycoprotein antigens used for serological diagnosis of NCC and the detection of other glycoconjugates was explored in cysticerci and the surrounding porcine tissue to understand their potential role in pathogenesis. Immunohistochemistry with an antiserum specific for glycoprotein antigens rich in N-linked carbohydrates and in situ histochemistry with a battery of lectins that have affinity to a variety of glycoconjugates were performed. The glycoconjugates rich in N-linked carbohydrates were detected in the vesicular fluid and tegument of the vesicular membrane and scolex, where the parasite has direct contact with the host tissues during cysticercosis and taeniasis, respectively. Additionally, as the inflammatory response progressed, the parasite's antigenic glycoproteins were also detected in the cytoplasm of inflammatory cells in the surrounding granuloma. In contrast, the spiral canal tegument, which will be exposed to intestinal enzymes in taeniasis, had N-acetyl-galactosamine-rich mucins. Thus, the differential saccharidic composition in T. solium metacestode structures may be important for the survival of the parasite in different host sites.     

Neurocysticercosis in Mexico

Cysticercosis is caused by the establishment of Taenia solium larvae (cysticerci), mainly in the central nervous system (CNS) and skeletal muscle of humans and pigs, after ingestion of eggs shed in human faeces by the adult tapeworm (see centrepage diagram). Human neurocysticercosis - often a life-threatening disease - is increasingly recognized as a public health problem, especially in developing countries. Clinical incidence of neurocysticercosis can reach 7% in Mexico and 18% in the Ekari population of New Guinea, while prevalence in autopsies ranges from 0.4% to 3.6% in several countries of Latin America, Asia and Africa. Many cases have also been recently reported in the USA, usually in immigrants. In this review, Ana Flisser focuses on the problems of cysticercosis in Mexico, where the disease is now recognized as a priority both in public health and economic terms. Recognition of the problem has been greatly aided in recent years by new developments in diagnosis - especially computed tomography (CT) to diagnose early stages of neurocysticercosis - and by improved drug treatment.     

Differential expression of calreticulin in developmental stages of Taenia solium

Taenia solium, a cestode that causes neurocysticercosis and taeniasis in humans, has a complex life cycle. The adult tapeworm develops in the intestine of human beings and is also responsible for neurocysticercosis, which is caused by the metacestode or cysticercus that develops in the brain. Recently, we have cloned the coding region for T. solium calreticulin (TsCRT) as a functional Ca(2+)-binding protein. Calreticulin is a ubiquitous protein involved in cellular Ca2+ homeostasis and protein folding. These important functions affect several aspects of cell physiology. To explore the expression of TsCRT during the T. solium life cycle, we used a specific polyclonal antibody raised against recombinant TsCRT to localize this protein by immunolabeling techniques. In sections of cysticerci obtained from swine muscle, as well as of adult tapeworms obtained after infection of hamsters with cysticerci, TsCRT was preferentially localized in tegumentary and muscle cytons of the suckers and rostellum. In mature proglottids obtained from infected humans, positive staining was observed in spermatogonia, ovogonia, uterine epithelium, and cells of the vas deferens. In the gravid uterus, the morula and early stage embryos were highly positive to TsCRT. However, expression diminished as embryonic development progressed and was absent in fully developed oncospheres that were surrounded by an embryophore. A similar down regulation was observed during spermatogenesis. Although early spermatocytes showed a high expression of TsCRT, mature spermatozoa present in the vas deferens were completely negative. These data indicate that calreticulin expression is spatially and temporally regulated during development of T. solium, especially during germ cell development and embryogenesis. In addition, these original images illustrate, for the first time, these processes at a histological level.     

Experimental infection of Philippine Taenia in domestic animals.

In the present study, six 34-44-day-old Small-Ear-Miniature pigs and one 14-day-old Holstein calf were each fed 10,000 Philippine Taenia eggs and sacrificed 27-43 days after inoculation. The infection rate was 100% for both pigs and calf with cysticerci recovery rates of 11 and 6%, respectively. A total of 6431 cysticerci were recovered only from the livers of the six pigs and 597 only from the liver of the calf; more occurred in the parenchyma (pigs 75%, calf 83%) than on the surface (pigs 25%, calf 17%). Mature cysticerci were found in four of the six pigs. A total of 317 cysticerci recovered from the pig livers were mature and the rest were either immature (926), degenerate or calcified (5188). All 597 cysticerci recovered from the liver of the calf were degenerate or calcified. Measurements of length, width, diameter of protoscolex, rostellum, and sucker and hooklet pattern indicated that Philippine Taenia is very similar to Taenia from Taiwan, Korea, Indonesia and Thailand and very different from classical T. saginata and T. solium.     

Host-cell apoptosis in Taenia solium-induced brain granulomas in naturally infected pigs

To assess whether apoptosis occurs in pig brain granulomas due to Taenia solium cysticerci, brain tissues from 30 pigs naturally infected with T. solium cysticercosis were evaluated by terminal deoxynucleotidyl transferase-end labelling (TUNEL) staining. In addition, tissues were stained with CD3 marker to identify T lymphocytes. Examination of TUNEL-stained tissues showed apoptotic cells in early lesions that contained viable cysticerci. Apoptotic cells were primarily found interspersed with normal cell types, and were mostly located in the inflammatory infiltrate. Late or advanced granulomas with disintegrated scolices did not show TUNEL-positive cells. CD3+ cells were found in both early and advanced lesions and apoptosis mainly co-localized with CD3+ T lymphocytes. This suggests that these cells are constantly undergoing apoptosis and thus die as soon as they arrive at the site of infection. Apoptosis indeed may be one way by which T. solium cysticerci down-regulate the host's cellular immune response in early cysticercosis. Therefore, further research is needed to establish if other cells besides T-lymphocytes are also a target for destruction by cysticerci in early cysticercosis as well as studies to assess if cysteine protease is expressed by viable cysticerci in situ.     

Experimental infection of Indonesia Taenia (Samosir strain) in domestic animals

Three 58-day old Small-Ear-Miniature (SEM) pigs, six 36-day old Landrace-Small-Ear-Miniature (L-SEM) pigs, and two 5-day old Holstein calves were each fed 3000 or 30,000 Indonesia Taenia (Samosir strain) eggs and sacrificed 27-195 days after inoculation. A total of 4922 cysticerci were recovered only from the livers of the three SEM pigs (1977 cysticerci) and six L-SEM pigs (2945 cysticerci). The infection rate in pigs was 100%. Cysticerci recovery rates of SEM and L-SEM pigs were 22.0% and 1.6%, respectively. Calves were not susceptible to Indonesia Taenia. More cysticerci were found in the liver parenchyma (L-SEM, 66.4%; SEM, 76.2%) than on the liver surface (L-SEM, 33.6%; SEM, 23.8%) of the infected animals. Most (99.86%) of the cysticerci recovered from the livers of L-SEM pigs were degenerated or calcified but 77.9% of those in the livers of SEM pigs were mature and only 22.1% were degenerated or calcified. Measurements of length, width, diameters of protoscolex, rostellum, and sucker and hooklet pattern indicated that Indonesia Taenia is very similar to Taiwan Taenia and very different from Taenia saginata and Taenia solium. The present findings indicate that Indonesia Taenia and Taiwan Taenia may be the same new species.     

The pig as an intermediate host for Taiwan Taenia infection

Eggs (1000-100,000/animal) of Taiwan Taenia were inoculated per os into 14 Small-Ear-Miniature (SEM), 19 Landrace-Small-Ear-Miniature (L-SEM), and 5 Duroc-Yorkshire-Landrace (DYL) pigs. These animals were sacrificed 7-107 days after infection. Thirty-four pigs were found to be infected with Taiwan Taenia cysticerci and the infection rates of SEM, L-SEM, and DYL were 86%, 89% and 100% respectively. The cysticerci recovery rates of SEM, L-SEM and DYL pigs were 27.2%, 1.7% and 0.27% respectively. Cysticerci were recovered only from the livers and none were found in muscles, viscera or other parts of the carcasses. More cysticerci were located in the liver parenchyma (71%) than on the liver surface (29%). Taiwan Taenia cysticerci were smaller than those of classical T. saginata or T. solium. Moreover, Taiwan Taenia cysticerci had 2 rows of rudimentary hooklets on the scolex. The results of this study indicate that young pigs are good intermediate hosts for Taiwan Taenia and that the SEM pig is a satisfactory host for experimental studies with this tapeworm. These results were similar to other studies with different geographic strains of the T. saginata-like tapeworm in the Far East. These strains appear to be the same and possibly a new species.     

Dogs as alternative intermediate hosts of Taenia solium in Papua (Irian Jaya), Indonesia confirmed by highly specific ELISA and immunoblot using native and recombinant antigens and mitochondrial DNA analysis

Serology (ELISA and immunoblot) using native glycoproteins, affinity purified glycoproteins, and a recombinant antigen is known to be highly specific to Taenia solium cysticercosis in humans and pigs. These techniques were applied for dogs in the highly endemic area of cysticercosis in Papua (Irian Jaya), Indonesia. Analysis of dog sera by both ELISA and immunoblot revealed 7 of 64 dogs were highly positive. Examination of two sero-positive dogs revealed cysticerci of T. solium in the brain and heart of these dogs. Mitochondrial DNA analysis confirmed that they were the same as T. solium previously confirmed from pigs and biopsies from local people from Irian Jaya. It is suggested that the life cycle of T. solium may be completed not only between humans and pigs but also between humans and dogs.     

The Asian Taenia and the possibility of cysticercosis

In certain Asian countries, a third form of human Taenia, also known as the Asian Taenia, has been discovered. This Asian Taenia seems to be an intermediate between Taenia solium and T. saginata since in morphological terms it is similar to T. saginata, yet biologically, as it uses the same intermediate host (pigs), it is more akin to T. solium. Taenia solium causes human cysticercosis, while T. saginata does not. It is not known whether the Asian taeniid is able to develop to the larval stage in humans or not. The arguments proposed by those authors who consider it unlikely that the Asian Taenia causes human cysticercosis are: (a) its molecular similarities with T. saginata; (b) the absence of cases of human cysticercosis in populations where the Asian adult is highly prevalent; and (c) the unsupporting results derived from an experimental infestation study. These three arguments are debated, although bearing in mind that at present there is still no clear scientific data to support that human cysticercosis can be caused by the Asian Taenia.     

Experimental infection of Thailand Taenia (Chiengmai strain) in domestic animals

Twenty-five gravid proglottides of a Thailand Taenia were obtained from a patient in Chiengmai, Thailand, and brought to our laboratory. The tapeworm was determined to be T. saginata-like by counting uterine branches (mean number 16, range 12-19 on each side). The eggs from these proglottides remained infective under storage at room temperature for 14 days followed by refrigeration (4-8 degrees C) for 131 days. Eight Small-Ear-Miniature pigs and two Holstein calves were each fed with 3000 eggs and sacrificed 12-76 days afterwards. Six pigs became infected and 16 cysticerci were recovered from the livers. Thirteen degenerated/calcified cysticerci were also recovered from the livers of the two calves. More cysticerci were found in the liver parenchyma (55%) than on the liver surface (45%) of the infected animals. Measurements of length, width, diameters of protoscolex, rostellum and sucker and hooklet pattern show that Thailand Taenia is similar to Taenia from Taiwan, Korea and Indonesia but different from T. saginata and T. solium. These findings indicate that Thailand Taenia, Taiwan Taenia, Korea Taenia, and Indonesia Taenia may be of the same species or sub-species.     

Protein and antigen diversity in the vesicular fluid of Taenia solium cysticerci dissected from naturally infected pigs

Cysticercosis caused by Taenia solium is a health threat for humans and pigs living in developing countries, for which there is neither a flawless immunodiagnostic test nor a totally effective vaccine. Suspecting of individual diversity of hosts and parasites as possible sources of the variations of the parasite loads among cysticercotic animals and of the limited success of such immunological applications as well as, we explored and measured both in nine cases of naturally acquired porcine cysticercosis. For this purpose, 2-Dimensional IgG immunoblots were performed by reacting the sera of each cysticercotic pig with the antigens contained in the vesicular fluid (VF) of their own cysticerci. We found an unexpectedly large diversity among the proteins and antigens contained in each of the nine VFs. Also diverse were the serum IgG antibody responses of the nine pigs, as none of their 2D- immunoblot images exhibited the same number of spots and resembled each other in only 6.3% to 65.3% of their features. So large an individual immunological diversity of the cysticercal antigens and of the infected pigs′ IgG antibody response should be taken into account in the design of immunological tools for diagnosis and prevention of cysticercosis and should also be considered as a possibly significant source of diversity in Taenia solium′s infectiveness and pathogenicity.     

Progesterone induces mucosal immunity in a rodent model of human taeniosis by Taenia solium

More than one quarter of human world's population is exposed to intestinal helminth parasites. The Taenia solium tapeworm carrier is the main risk factor in the transmission of both human neurocysticercosis and porcine cysticercosis. Sex steroids play an important role during T. solium infection, particularly progesterone has been proposed as a key immunomodulatory hormone involved in susceptibility to human taeniosis in woman and cysticercosis in pregnant pigs. Thus, we evaluated the effect of progesterone administration upon the experimental taeniosis in golden hamsters (Mesocricetus auratus). Intact female adult hamsters were randomly divided into 3 groups: progesterone-subcutaneously treated; olive oil-treated as the vehicle group; and untreated controls. Animals were treated every other day during 4 weeks. After 2 weeks of treatment, all hamsters were orally infected with 4 viable T. solium cysticerci. After 2 weeks post infection, progesterone-treated hamsters showed reduction in adult worm recovery by 80%, compared to both vehicle-treated and non-manipulated infected animals. In contrast to control and vehicle groups, progesterone treatment diminished tapeworm length by 75% and increased proliferation rate of leukocytes from spleen and mesenteric lymph nodes of infected hamsters by 5-fold. The latter exhibited high expression levels of IL-4, IL-6 and TNF-α at the duodenal mucosa, accompanied with polymorphonuclear leukocytes infiltration. These results support that progesterone protects hamsters from the T. solium adult tapeworm establishment by improving the intestinal mucosal immunity, suggesting a potential use of analogues of this hormone as novel inductors of the gut immune response against intestinal helminth infections and probably other bowel-related disorders.     

In vitro hydroxylation of proline in the collagen of the cysticercus of Taenia solium

1. In vitro hydroxylation of proline in cysticercus collagen was carried out using two different vertebrate enzymes. 2. Chick embryo enzyme is more active on cysticercus collagen than new-born rabbit enzyme. 3. Hydroxylation of cysticercus collagen is more efficient at 40 than at 37 degrees C. 4. No prolyl hydroxylase activity was detected in cysticercus extracts. 5. Collagen from the adult tapeworms Taenia solium and T. saginata lack hydroxyproline. 6. SLS crystallites from T. solium and T. saginata collagen show the same band pattern as cysticercus collagen.     

A novel,sequencing-free strategy for the functional characterization of Taenia solium proteomic fingerprint

The flatworm Taenia solium causes human and pig cysticercosis. When cysticerci are established in the human central nervous system, they cause neurocysticercosis, a potentially fatal disease. Neurocysticercosis is a persisting public health problem in rural regions of Mexico and other developing countries of Latin America, Asia, and Africa, where the infection is endemic. The great variability observed in the phenotypic and genotypic traits of cysticerci result in a great heterogeneity in the patterns of molecules secreted by them within their host.This work is aimed to identify and characterize cysticercal secretion proteins of T. solium cysticerci obtained from 5 naturally infected pigs from Guerrero, Mexico, using 2D-PAGE proteomic analysis. The isoelectric point (IP) and molecular weight (MW) of the spots were identified using the software ImageMaster 2D Platinum v.7.0. Since most secreted proteins are impossible to identify by mass spectrometry (MS) due to their low concentration in the sample, a novel strategy to predict their sequence was applied. In total, 108 conserved and 186 differential proteins were identified in five cysticercus cultures. Interestingly, we predicted the sequence of 14 proteins that were common in four out of five cysticercus cultures, which could be used to design vaccines or diagnostic methods for neurocysticercosis. A functional characterization of all sequences was performed using the algorithms SecretomeP, SignalP, and BlastKOALA. We found a possible link between signal transduction pathways in parasite cells and human cancer due to deregulation in signal transduction pathways. Bioinformatics analysis also demonstrated that the parasite release proteins by an exosome-like mechanism, which could be of biological interest.     

Proteomic analysis of Taenia solium metacestode excretion-secretion proteins

The metacestode larval stage of Taenia solium is the causal agent of a zoonotic disease called cysticercosis. The disease has an important impact on pork trade (due to porcine cysticercosis) and public health (due to human neurocysticercosis). In order to improve the current diagnostic tools and to get a better understanding of the interaction between T. solium metacestodes and their host, there is a need for more information about the proteins that are released by the parasite. In this study, we used protein sequences from different helminths, 1DE, reversed-phase LC, and MS/MS to analyze the excretion-secretion proteins produced by T. solium metacestodes from infected pigs. This is the first report of the T. solium metacestode excretion-secretion proteome. We report 76 proteins including 27 already described T. solium proteins, 17 host proteins and 32 proteins likely to be of T. solium origin, but identified using sequences from other helminths.     

Spatial distribution of Taenia solium porcine cysticercosis within a rural area of Mexico

Cysticercosis is caused by Taenia solium, a parasitic disease that affects humans and rurally bred pigs in developing countries. The cysticercus may localize in the central nervous system of the human, causing neurocysticercosis, the most severe and frequent form of the disease. There appears to be an association between the prevalence of porcine cysticercosis and domestic pigs that wander freely and have access to human feces. In order to assess whether the risk of cysticercosis infection is clustered or widely dispersed in a limited rural area, a spatial analysis of rural porcine cysticercosis was applied to 13 villages of the Sierra de Huautla in Central Mexico. Clustering of cases in specific households would indicate tapeworm carriers in the vicinity, whereas their dispersal would suggest that the ambulatory habits of both humans and pigs contribute to the spread of cysticercosis. A total of 562 pigs were included in this study (August-December 2003). A global positioning system was employed in order to plot the geographic distribution of both cysticercotic pigs and risk factors for infection within the villages. Prevalence of pig tongue cysticercosis varied significantly in sampled villages (p = 0.003), ranging from 0% to 33.3% and averaging 13.3%. Pigs were clustered in households, but no differences in the clustering of cysticercotic and healthy pigs were found. In contrast, the presence of pigs roaming freely and drinking stagnant water correlated significantly with porcine cysticercosis (p = 0.07), as did the absence of latrines (p = 0.0008). High prevalence of porcine cysticercosis proves that transmission is still quite common in rural Mexico. The lack of significant differentiation in the geographical clustering of healthy and cysticercotic pigs weakens the argument that focal factors (e.g., household location of putative tapeworm carriers) play an important role in increasing the risk of cysticercosis transmission in pigs. Instead, it would appear that other wide-ranging biological, physical, and cultural factors determine the geographic spread of the disease. Extensive geographic dispersal of the risk of cysticercosis makes it imperative that control measures be applied indiscriminately to all pigs and humans living in this endemic area.