Sparganosis: an under-recognised zoonosis in Australia?

Sparganosis is a rare zoonotic parasitosis that is sporadically reported worldwide. In Australia, the causative tapeworms are considered endemic in wildlife animals, however, there have been only five reported human infections. We present three additional cases of sparganosis, involving two Australian born gentlemen who have never travelled overseas and a woman who emigrated from Ethiopia. The first man presented with two unusual subcutaneous lumps that migrated along the anterior abdominal wall connected by a tunnel. The second man presented with two separate lumps, one on the thigh and the other on the left upper abdomen over a 4-week interval. The woman presented with 6 weeks of intermittent fevers, night sweats, abdominal pain and passing intestinal worms. This series of patients suggests that sparganosis is under-recognised in Australia and serves as a reminder for clinicians to the varied presentations that can be characteristic of this lesser known zoonosis.

Establishment of the complete life cycle of Spirometra (Cestoda: Diphyllobothriidae) in the laboratory using a newly isolated triploid clone.

Methods to maintain the life cycle of pathogenic organisms become powerful tools for studying molecular and cellular bases of infectious diseases. Spirometra erinaceieuropaei is a parasitic tapeworm that causes sparganosis in humans. Because S. erinaceieuropaei has a complex life cycle with different stages and host species requirements, there have been no reports to establish the complete life cycle in the laboratory. In this study, using Cyclops as the first intermediate host, mouse as the experimental second intermediate host, and dog as the final host, we succeeded in maintaining S. erinaceieuropaei in the laboratory. By repeating the established life cycle five times, we obtained a clonal population of S. erinaceieuropaei from a single adult worm. A karyotype study showed that the chromosome of this clone is triploid (3n=27), indicating that a genetically uniform strain is established by apomictic reproduction. The strain was named Kawasaki triploid (Kt). A partial sequence of mitochondrial cytochrome c oxidase subunit 1 gene of the strain Kt showed more than 98% similarity with those of S. erinaceieuropaei isolates from Australia, China, and South Korea, and the resultant phylogeny indicated that the strain Kt is a member of a distinctive clade from East Asia and Oceania. Our system will be particularly useful for studies of S. erinaceieuropaei infection and human sparganosis.

The life-cycle of Spirometra species from Peninsular Malaysia.

The life-cycle of Malaysian Spirometra spp. was studied under experimental conditions in the laboratory. The Cyclops were reared as the first intermediate host, the hamster as the experimental second intermediate host and cat as the definitive host. Maturation and hatching of eggs took 6 to 12 days by incubation at temperature 30 ºC. The hatched coracidium measured 46 x 34 µm. The Cyclops used were susceptible to the coracidial infection. The procercoid older than 5 days in the Cyclop body cavity had minute spines at the anterior end, calcium corpuscles in the body parenchyma and the cercomer at the posterior end. Procercoids 10 to 14 days old were infective to hamster. The plerocercoids from the hamster after 30 days were long and slender and were infective to cats. The plerocercoids experimentally inoculated to cats developed to adult worms and began to produce eggs between 10 to 60 days. Based on the results that have been obtained, a complete life-cycle was successfully elucidated in the laboratory and hamster was identified to be a good laboratory model for a second intermediate host of Spirometra sp.

The Pampas fox (Lycalopex gymnocercus) as new definitive host for Spirometra erinacei (Cestoda: Diphyllobothriidae).

Spirometra erinacei, Faust, Campbell and Kellogg, 1929, is a pseudophyllidean cestode of the family Diphyllobothriidae. The genus Spirometra is cosmopolitan and these parasites infect carnivores, specially felids and canids. In Argentina, S. erinacei and S. mansonoides have been reported sporadically only in domestic definitive hosts. The Pampas fox, Lycalopex gymnocercus, is the most abundant native carnivore in southern South America, where it inhabits grasslands and open woodlands and areas highly modified by extensive ranching and agricultural activities. This report describes the first finding of S. erinacei infecting Pampas fox, and provides an estimate prevalence of this cestode in rural areas of southern Buenos Aires province, Argentina based on 78 complete Pampas fox intestine samples analysis. This study found a 15.4% of prevalence of S. erinacei in small intestine (adult stage) and a 21.8% in fecal samples (egg stage). In the present work, the first case of S. erinacei in a wild definitive host from Argentina was reported expanding the list of definitive hosts of S. erinacei in South America.

MR spectroscopy and MR perfusion character of cerebral sparganosis: a case report.

The authors report the case of a 46-year-old woman with cerebral sparganosis resulting from infection with a larva of Spirometra. Computed tomography and magnetic resonance imaging revealed a mass lesion with prominent perifocal oedema in the left parietal lobe. Advanced imaging pulse sequences, including MR spectroscopy and MR perfusion, were performed. During surgery for the removal of a granuloma, the parasite was discovered and excised. Following treatment, the patient's neurological deficits markedly improved.

Differential protein expression in Spirometra erinacei according to its development in its final host.

This study was undertaken to identify genes involved in the growth and development of Spirometra erinacei larvae, an intestinal tapeworm of cats and dogs, within the final host. The differential protein expression at three different stages of S. erinacei, the plerocercoid larvae, 8-day-old juveniles, and adults, was compared using two-dimensional electrophoresis. Specifically or highly expressed proteins in juvenile worms were analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometer (MS)/MS. The proteome map of larvae showed fewer protein spots than juveniles or adults, whereas juveniles or adults revealed a similar protein expression profile. Eight juvenile-specific and five upregulated proteins of juveniles were identified and matched to proteins of known biological functions. These were grouped into several categories of functionally related proteins: DNA/RNA metabolism, cell trafficking, cytoskeleton, protein processing and degradation, energy metabolism, and oxidative stress. Our results give an overview of the growth and development mechanisms of cestodes within the final host and extend our understanding of parasite biology in the host-parasite relationship.

Neuroimaging and pathological findings in a child with cerebral sparganosis. Case report.

The authors report the case of a 6-year-old boy with cerebral sparganosis due to infection with a plerocercoid tapeworm larva of Spirometra mansoni. Magnetic resonance imaging revealed an area of irregular long T2 signal in the right frontal lobe. When compared with images obtained 2 years earlier, the lesion appeared to have migrated into the parietal lobe. During surgery for the removal of a granuloma, the parasite was discovered and excised. Following surgery, the patient's neurological deficits markedly improved. The authors review the pathological and imaging features of cerebral sparganosis.

Cloning of the novel putative apoptosis-related gene of Spirometra erinacei (Order Pseudophyllidae).

We postulated that apolysis was processed in accordance with apoptotic changes occurring in a cestode, Spirometra erinacei (Pseudophyllidea). We cloned the novel putative apoptosis-associated gene from S. erinacei via screening of a S. erinacei cDNA library with a ced-3 gene (activator of apoptosis) probe from Caenorhabditis elegans. We identified a 261-bp cDNA sequence, which encodes for an 86-amino acid protein. The cloned gene expression was observed in the neck and gravid proglottids via Northern blotting, using cloned cDNA inserts as probes, but the clone was not expressed in any of other tissues. We suggest that this gene may be involved in the apolysis of S. erinacei during normal tissue development and differentiation in cestode parasites.

A review of human sparganosis in Thailand.

BACKGROUND: Sparganosis is a zoonosis that occurs occasionally in humans. The infection is reported in many countries but is most common in eastern Asia. In Thailand, a southeast Asian country, the infection is sporadic. DESIGN: In this study the clinical presentations of human sparganosis cases in Thailand were investigated by means of a literature review. RESULTS: Reports of 34 cases of sparganosis were found. The infections were ocular (17 cases), subcutaneous (ten cases), central nervous system (five cases), auricular (one case), pulmonary (one case), intraosseous (one case) and intraperitoneal (one case). Of these 34 cases, 14 had risk behaviour reported, 12 had a history of drinking impure water, five had a history of eating frog or snake meat and two had a history of using frog or snake meat as a poultice. Some cases had more than one risk factor. CONCLUSION: Most cases of sparganosis in Thailand presented with superficial ocular mass lesions. The major risk behaviour in Thailand is drinking water contaminated with the infective organism. Some cases of serious deep visceral sparganosis have also been reported.

Purification and localization of a 10 kDa calcareous corpuscle binding protein of Spirometra mansoni plerocercoid.

In cestode parasites, calcareous corpuscles are thought to be associated with a number of intracellular physiologies by regulating the trafficking of mineral components. We previously separated these particular components by Ficoll, and their binding proteins of 10 kDa and 35 kDa in the Spirometra mansoni plerocercoid (sparganum). In the present study, we purified a 10 kDa protein employing octyl-Sepharose CL-4B affinity chromatography. Anti-serum raised against the purified protein showed specific reactions to the calcareous corpuscles of the worm section by immunohistochemistry, and recognized the 10 kDa protein by immunoblotting.

Cestode parasites: application of in vivo and in vitro models for studies on the host-parasite relationship.

Cestode worms, commonly also known as 'flat' worms or tapeworms, are an important class of endoparasitic organisms. In order to complete their life cycle, they infect intermediate and definitive hosts in succession, through oral ingestion of eggs or larvae, respectively. Serious disease in humans or other mammalian hosts is mostly caused by the larval stages. Echinococcus spp. and Taenia spp. have been extensively investigated in the laboratory due to the fact that they represent important veterinary medical challenges and also cause grave diseases in humans. In contrast, Hymenolepis spp. and Mesocestoides spp. infections are relatively rare in humans, but these parasites have been extensively studied because their life cycle stages can be easily cultured in vitro, and can also be conveniently maintained in laboratory animal hosts. Thus they are more easily experimentally accessible, and represent important models for investigating the various aspects of cestode biology. This review will focus on in vitro and in vivo models which have been developed for studies on the host-parasite relationship during infection with Echinococcus, Taenia, Hymenolepis, Mesocestoides and Spirometra, and will cover the use of these models to investigate the morphology and ultrastructure of respective genera, the immunological relationship with the host and the development of vaccination approaches, as well as applications of these models for studies on parasite metabolism, physiology and gene expression. In addition, the use of these models in the development of chemotherapeutic measures against cestode infections is reviewed.

Differential expression of the 27 kDa cathepsin L-like cysteine protease in developmental stages of Spirometra erinacei.

The 27 kDa cathepsin L-like cysteine protease of Spirometra erinacei plerocercoid is known to play an important function in tissue penetration, nutrient uptake and immune modulation in human sparganosis. In the present study, the expression of this enzyme was examined at different developmental stages of S. erinacei including immature egg, coracidium, plerocercoid in tadpole and rat, and adult. Proteolytic activity against carboxybenzoyl-phenylalanyl-arginyl-7-amino-4-methylcoumarin was detected in the extracts of coracidia and plerocercoid while no activity was observed in those of immature egg and adult. The specific activity in coracidial extracts was lower than that in the plerocercoid. Reverse transcription-polymerase chain reaction and Northern blot analysis demonstrated that the gene was expressed in the coracidium and plerocercoid but not in immature egg and adult. These results suggest that the 27 kDa cysteine protease is only expressed in the stages involving active migration of the parasite in the host tissue.

A monoclonal antibody against a glycolipid SEGLx from Spirometra erinaceieuropaei plerocercoid.

A mouse monoclonal antibody AK97 (IgM) was established against a new type of glycosphingolipid, SEGLx, isolated from plerocercoids of tapeworm, Spirometra erinaceieuropaei. The chemical structure of SEGLx (Gal beta1-4(Fuc alpha1-3)(Glc beta1-3Gal beta1-ceramide) had been previously characterized. The specificity of AK97 was determined by thin-layer chromatography-immunostaining and enzyme-linked immunosorbent assay. AK97 was found to be directed to SEGLx and GalSEGLx (Gal beta1-4(Fuc alpha1-3)Glc beta1-3(Gal beta1-6)Gal beta1-ceramide) and also showed cross-reactivity with the stage specific embryonic antigen-1 (SSEA-1), the epitope being defined to be the non-reducing terminal trisaccharide sequence. On immunohistochemical examination, AK97 predominantly stained the tegument, the external surfaces of worms which have a brush border-like organization. Based on the immunohistochemical findings for the staining liability as to organic solvents and the results of Western blot analysis of the plerocercoid glycoproteins, it was proved that the antigens in the tapeworm were glycolipids. Considering that the tapeworm is in direct contact with its host's tissue through the tegument, the membrane surface of which is exposed to the external environment, it is suspected that SEGLx and GalSEGLx on the tegument play functionally important roles in the host parasite interaction.

Excretory/secretory products of plerocercoids of Spirometra erinaceieuropaei induce the expression of inducible nitric oxide synthase mRNA in murine hepatocytes.

In this study, we observed the level of normal murine hepatocyte inducible NOS (iNOS) mRNA by semi-quantitative polymerase chain reaction (SQ-PCR) analysis after stimulation with ES products (ESP) and/or ESP fractions from the plerocercoids. We found that ESP are able to induce the expression of iNOS gene in a dose-dependent fashion. Treatment of ESP with polymyxin B did not affect their ability to induce the expression of iNOS gene, suggesting that bacterial lipopolysaccharide (LPS) is not involved. The iNOS-inducing factor (a) is soluble, and may be a component whose molecular mass exceeds 94 kDa as analyzed by a combination of SDS-PAGE and SQ-PCR. The peak of iNOS mRNA level was detected 3 h after stimulation with ESP; the mRNA level decreased sharply from 9 h. Dexamethasone inhibited the induction of mRNA for hepatocyte iNOS. In contrast, cycloheximide stimulated the induction; this suggests that de nova protein synthesis is important in the regulation of the ESP-induced expression of iNOS mRNA. Actinomycin D blocked the induction. In addition, the results of Northern blot analysis showed that ESP suppressed the LPS (10 micrograms/ml) and interferon-gamma (IFN-gamma, 100 U/ml)-induced hepatocyte iNOS mRNA expression in a dose-dependent fashion and the suppressing effect was more marked when hepatocytes were exposed to ESP 3 h prior to LPS and IFN-gamma. These results demonstrate that the soluble factor(s) of ESP is capable of inducing murine iNOS gene expression in hepatocytes in the absence of added cytokines.

Monohexosylceramides of larval and adult forms of the tapeworm, Spirometra erinacei.

The occurrence of glycosphingolipids with unique carbohydrate structures in different species of cestode, Platyhelminth, which had been shown previously, prompted us to study the molecular species of the monohexosylceramides (cerebrosides) in the pseudophyllidean cestode, Spirometra erinacei. The purpose of the study was to obtain a basis for future investigations of the physiological role of glycolipids in parasitism. Cerebrosides were isolated from S. erinacei at two growth stages, i.e., from the larval form (plerocercoid) and from the adult tapeworms (intestinal form). The cerebrosides were separated into four subfractions by silica gel column chromatography, and their constituents were analyzed by gas-liquid chromatography, gas chromatography/mass spectrometry, and high-performance liquid chromatography/mass spectrometry. The hexoses of the cerebrosides consisted primarily of galactose in both growth stages, while only a small amount of glucose was detected. The ceramides were composed of sphinganine (d18:0) and phytosphingosine (t18:0) as sphingoid bases, and of nonhydroxy fatty acids ranging from C16 to C30 and hydroxy stearic acid (18h:0). The cerebrosides of adult tapeworms contained more 18h:0 than those of plerocercoids. The combination of hexoses and ceramides in the cerebroside molecules was slightly different in the two growth stages: the glucocerebrosides of plerocercoids contained only d18:0-nonhydroxy fatty acids in their ceramide moieties, whereas those of adult tapeworms contained varying ceramide moieties. Our data indicate that the molecular species of glycolipids present were essentially homeostatic throughout growth in spite of the entirely different environmental conditions, although there were slight differences in the hexose distribution in the two growth stages.

The fatty acids of each lipid fraction and their use in providing energy source of the plerocercoid of Spirometra erinacei.

The fatty acid concentration of each lipid fraction of plerocercoids of Spirometra erinacei and the host snake serum was investigated. The major fatty acids of phospholipid of the plerocercoids were C18:1, C18:0 and C16:0, and those of the host snake serum were C16:0, C18:1 and C18:0, in order of amount in both cases. The changes of the fatty acid composition of phospholipid of the plerocercoids when they were incubated in physiological saline at 18 degrees C and at 37 degrees C for 24 h were investigated in both cases. Polyunsaturated fatty acids increased at 18 degrees C, and saturated fatty acids increased at 37 degrees C. Michaelis constants (Km) of beta-hydroxyacyl-CoA dehydrogenase (HAD), NADH: ubiquinone oxidoreductase (complex I) (NADH: ferricyanide reaction) and complex I (NADH: ubiquinone reaction) for NADH were 20.6, 50 and 13.3 microM, respectively. The ATP production in mitochondria of the plerocercoids was accelerated by adding ADP and inhibited by adding such electron transport system inhibitors as rotenone, antimycin A and sodium cyanide. These results suggested that the fatty acids in the plerocercoids played an important role in regulating the fluidity of membrane by changing the composition in membrane lipid corresponding with the change of temperature circumstance. The NADH reduced by HAD might be accepted by the complex I in the electron transport system, and thus the parasites were capable of ATP production in a classical pathway of the oxidative phosphorylation system.

Ultrastructure studies on the papillae and the nonciliated sensory receptors of adult Spirometra erinacei (Cestoda, Pseudophyllidea).

The small numerous papillae on the ventral surface of the gravid proglottid of adult Spirometra erinacei were studied by scanning electron microscopy. The arrangement of clumps of papillae was recognized on the surface of the central portion around the genital atrium, with lateral clumps being located above a pair of longitudinal nerve cords and marginal ones, on both sides of the proglottid. By transmission electron microscopy, two types of nonciliated sensory receptors were observed within the papillae. The type I, single receptor was embedded within a papilla. This dome-like sensory receptor contained two electron-dense collars and four rootlets surrounded by numerous thin filaments. The type II receptor was found arranged in groups in the area between the papillae, and the apical end was exposed to the external environment. This simple, club-like sensory receptor contained electron-lucent vesicles and microtubules. We believe that the papillae play an important role in cross-insemination.

Experimental life history of Spirometra erinacei.

The complete life cycle of Spirometra erinacei has been experimentally maintained in the laboratory. The cyclops were reared as the first intermediate host, and the tadpoles of Rana nigromaculata as the second intermediate host. ICR mice were used as another second host. The experimental definitive hosts were dogs and cats. Maturation and hatching of the eggs took 8 to 14 days by incubation at 29 degrees C. The coracidium measured 43.8 x 36.9 microns. Mesocyclops leuckarti and Eucyclops serrulatus were susceptible to the coracidial infection. The procercoids older than 5 days in the cyclops had minute spines at the anterior end, calcium corpuscles in the body parenchyme and the cercomer at the posterior end. Procercoids 10 to 20 days old were infective to tadpoles, and 15 or 21 day old worms could infect the mice. The plerocercoids from the tadpoles at 15 days after experimental infection were pear-shaped and shorter than 1 mm in the length and were infective to mice. Fifteen to 18 days after experimental inoculation of plerocercoids to dogs or cats, the adult worms began to produce eggs. One life cycle from egg to egg needed 48 to 67 days in the laboratory. The morphology of larval or adult worms was compatible with the description of Spirometra erinacei.

Significance of the distribution of 57Co-vitamin B12 in Spirometra mansonoides (Cestoidea) during growth and differentiation in mammalian intermediate and definitive hosts.

The distribution of labeled cyanocobalamin (CN-[57Co]Cbl = [57Co]-vitamin B12) in pleurocercoids and adult tapeworms of Spirometra mansonoides was studied during development in mice 22 days days PI, respectively. Plerocercoid scolices, obtained by cutting away their bodies or by in vitro enzymatic dissolution of the bodies, were pulsed with CN- magnitude of 57Co Cbl for 1h at 37 degrees C and reimplanted subcutaneously into mice or given per os to cats. In regenerated plerocercoids, the highest concentration of magnitude of 57Co Cbl occurred in the scolex and then decreased posteriorly in the newly-formed tissues of the body. Approximately 60% of the total magnitude of 57Co Cbl present remained concentrated in the scolex following body regeneration plerocercoids and adult tapeworms of Spirometra mansonoides was studied during development in mice 22 days post-infection (PI) and in cats 16 days PI, respectively. Plerocercoid scolices, obtained by cutting away their bodies or by in vitro enzymatic dissolution of the bodies, were pulsed with CN-[57Co]Cbl for 1 h at 37 degrees C and reimplanted subcutaneously into mice or given per os to cats. In regenerated plerocercoids, the highest concentration of [57Co]Cbl occurred in the scolex and then decreased posteriorly in the newly-formed tissues of the body. Approximately 60% of the total [57Co]Cbl present remained concentrated in the scolex following body regeneration for up to 109 days PI. This high [57Co]Cbl concentration in the plerocercoid scolex was bound to protein and appears to be maintained by a complex homeostatic mechanism in association with directional transport of [57Co]Cbl to the scolex with ultimate depletion along the length of the body.(ABSTRACT TRUNCATED AT 250 WORDS)