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Purpose: The aim of this study is to conduct a molecular characterization of Spirometra tapeworm from jungle cat (Felis chaus) in Guilan Province, north of Iran using DNA sequence analysis of the mitochondrial cytochrome c oxidase subunit 1 (Cox1) and 12S rDNA sequences. Methods: Morphological features of the adult tapeworm of Spirometra were evaluated using specific staining and light microscopy. The molecular characterization was performed using partial Cox1 and 12S rDNA regions. Genetic diversity was calculated and phylogenetic trees of the obtained sequences were constructed. Results: Morphological features were compatible with previous description of adult Spirometra erinaceieuropaei. The Cox1 sequence of the specimen showed 100% similarity with S. erinaceieuropaei sequences in GenBank from Korea, China and Iran. Also, the 12S rDNA sequence revealed 99.7% similarity with S. erinaceieuropaei isolates from China and Japan. Intra-species variation within isolates of S. erinaceieuropaei was 0–1.4% and 0–4.6% for Cox1 and 12S rDNA genes, respectively. Conclusion: This is the first report of molecular characterization of S. erinaceieuropaei in jungle cat, F. chaus in Iran. Jungle cat probably plays a major role as reservoir host in maintaining of this parasite in this area with favorable climate condition. Needs for further assessment on the role of appropriate hosts, especially intermediate/paratenic hosts as well as the potential risk of human infectivity with sparganosis is emphasized. [ABSTRACT FROM AUTHOR]

AbstractCystatin plays a crucial role in immune evasion by parasites. It regulates diverse immune response processes, such as antigen presentation, cytokine and NO production, and phagocytosis. In recent years, an increasing number of parasite cystatins have been identified and studied for the treatment of inflammatory diseases. In contrast, cystatin from Spirometra erinaceieuropaei has received limited research attention. The objective of this study was to utilize bioinformatics tools and molecular biology techniques to predict the biological properties and obtain a recombinant cystatin analogue from Spirometra erinaceieuropaei (SeCystatin). The SeCystatin gene consists of 417 bp and encodes a putative 100-amino acid protein. The predicted molecular weight and isoelectric point of SeCystatin were 10.89 kDa and 6.82, respectively; SeCystatin possesses 12 phosphorylation sites and five post-translational modification sites but has no signal peptide and transmembrane region. The secondary structure of SeCystatin consists of one α-helix, four β-folds and six coils. It also possesses a cystatin-specific conserved domain, QxVxG, which is positioned at the boundary between the first and second β-folds, thereby demonstrating the characteristic features of type I cystatin. In the context of molecular evolution, SeCystatin demonstrates the highest level of evolutionary similarity to Schistocephalus solidus. The SeCystatin gene was fully synthesized, and the recombinant plasmid pET-30a (+)-SeCystatin was constructed to express the target protein in Escherichia coli BL 21. Recombinant SeCystatin was successfully induced for expression by IPTG and subsequently purified using affinity chromatography (Ni-IDA). Our study provides a solid foundation for further studies on the biological functions of SeCystatin.

Cystatin plays a crucial role in immune evasion by parasites. It regulates diverse immune response processes, such as antigen presentation, cytokine and NO production, and phagocytosis. In recent years, an increasing number of parasite cystatins have been identified and studied for the treatment of inflammatory diseases. In contrast, cystatin from Spirometra erinaceieuropaei has received limited research attention. The objective of this study was to utilize bioinformatics tools and molecular biology techniques to predict the biological properties and obtain a recombinant cystatin analogue from Spirometra erinaceieuropaei (SeCystatin). The SeCystatin gene consists of 417 bp and encodes a putative 100-amino acid protein. The predicted molecular weight and isoelectric point of SeCystatin were 10.89 kDa and 6.82, respectively; SeCystatin possesses 12 phosphorylation sites and five post-translational modification sites but has no signal peptide and transmembrane region. The secondary structure of SeCystatin consists of one α-helix, four β-folds and six coils. It also possesses a cystatin-specific conserved domain, QxVxG, which is positioned at the boundary between the first and second β-folds, thereby demonstrating the characteristic features of type I cystatin. In the context of molecular evolution, SeCystatin demonstrates the highest level of evolutionary similarity to Schistocephalus solidus. The SeCystatin gene was fully synthesized, and the recombinant plasmid pET-30a (+)-SeCystatin was constructed to express the target protein in Escherichia coli BL 21. Recombinant SeCystatin was successfully induced for expression by IPTG and subsequently purified using affinity chromatography (Ni-IDA). Our study provides a solid foundation for further studies on the biological functions of SeCystatin. [ABSTRACT FROM AUTHOR]

Background: Sparganosis is a zoonotic disease caused by Plerocercoid larvae (spargana) of the genus Spirometra. We aimed to provide molecular evidence for the infection of amphibians with Spirometra sp. in the inside and outside of Horton Plains National Park (HPNP), Sri Lanka. Methods: The prevalence of sparganum infection in wild frogs (Truga eques and Minverya agricola) was investigated in the inside and outside of HPNP from June 2019 to April 2021.A total of 1,434 Amphibians samples were surveyed to examine the spargana infection from the study site. To identify the species identity of the collected spargana, a portion of the mitochondrial cytochrome c oxidase subunit 1 (cox1) gene and nuclear 18S rRNA gene were amplified, sequenced, and analyzed. Results: A total of 539 infected amphibians (T. eques and M. agricola) samples were examined to survey from the study area. Spargana were detected in all dissected specimens belonging to the species Spirometra erinaceieuropaei that were genetically confirmed using the evolutionary conserved nuclear 18S rRNA gene and then compared to the GenBank deposit, indicating that S. erinaceieuropaei is the primary causal agent of sparganosis both inside and outside the HPNP. Conclusion: Our finding is the first genetically confirmed record of S. erinaceieuropaei in amphibians in South Asia. However, further studies are needed to investigate the prevalence of sparagna infection in amphibians all over the island.

Background: Sparganosis is a zoonotic disease caused by Plerocercoid larvae (spargana) of the genus Spirometra. We aimed to provide molecular evidence for the infection of amphibians with Spirometra sp. in the inside and outside of Horton Plains National Park (HPNP), Sri Lanka. Methods: The prevalence of sparganum infection in wild frogs (Truga eques and Minverya agricola) was investigated in the inside and outside of HPNP from June 2019 to April 2021.A total of 1,434 Amphibians samples were surveyed to examine the spargana infection from the study site. To identify the species identity of the collected spargana, a portion of the mitochondrial cytochrome c oxidase subunit 1 (cox1) gene and nuclear 18S rRNA gene were amplified, sequenced, and analyzed. Results: A total of 539 infected amphibians (T. eques and M. agricola) samples were examined to survey from the study area. Spargana were detected in all dissected specimens belonging to the species Spirometra erinaceieuropaei that were genetically confirmed using the evolutionary conserved nuclear 18S rRNA gene and then compared to the GenBank deposit, indicating that S. erinaceieuropaei is the primary causal agent of sparganosis both inside and outside the HPNP. Conclusion: Our finding is the first genetically confirmed record of S. erinaceieuropaei in amphibians in South Asia. However, further studies are needed to investigate the prevalence of sparagna infection in amphibians all over the island. [ABSTRACT FROM AUTHOR]

To clarify the characteristics of Spirometra erinaceieuropaei fatty acid-binding protein (SeFABP) and obtain its recombinant protein, the basic physio-biochemistry characteristics, signal peptides, antigen epitopes, transmembrane domains, secondary and tertiary structures, multi sequence alignment and molecular evolutionary tree of SeFABP were predicted and analyzed. On this basis, SeFABP was wholegenome synthesized and cloned into prokaryotic expression vector. The recombinant plasmid pET- 30a(+)-SeFABP was constructed, then transformed into E. coli BL21 (DE3) and induced by IPTG. The purified SeFABP was obtained by Ni-IDA resins affinity chromatography, and were finally confirmed by SDS-PAGE and Western blot. The results showed that SeFABP, a stable intracellular protein without signal peptide and transmembrane region, was composed of 130 amino acids (AA). It contained 10 phosphorylation sites and 7 post-translational modification sites. Both methods predicted that SeFABP contained 5 linear epitopes, and were highly coincident, indicating that SeFABP had good immunogenicity. Its secondary structure contains 2 a-helix (13.07%), 10 ß-sheet (56.92%) and 30% random coli. The comparison of multiple FABPs sequences showed that the species were identical and highly conserved at multiple AA sites. Specifically, SeFABP has the highest similarity with T. multiceps and M. vogae, close to 37%, but the lowest similarity with F. hepatica and F. gigatica, close to 22%. Plasmid double enzyme digestion and sequencing showed that the fully synthesized SeFABP fragment was accurately cloned and connected to the expression vector pET-30a(+)-SeFABP. After induction, the recombinant SeFABP was successfully expressed in the form of inclusion body. SDS-PAGE and Western blot showed a 15kD band, indicating that the recombinant was successfully purified. In conclusion, the characteristics of SeFABP and its high-purity recombinant protein were obtained, provides a basis for future research based on SeFABP in vaccine development and drug design. [ABSTRACT FROM AUTHOR]

Copyright of Chinese Journal of Preventive Veterinary Medicine / Zhongguo Yufang Shouyi Xuebao is the property of Chinese Journal of Preventive Veterinary Medicine Editorial Office and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Human sparganosis is a food-borne parasitic disease caused by the plerocercoids of Spirometra species. Clinical diagnosis of sparganosis is crucial for effective treatment, thus it is important to identify sensitive and specific antigens of plerocercoids. The aim of the current study was to identify and characterize the immunogenic proteins of Spirometra erinaceieuropaei plerocercoids that were recognized by patient sera. Crude soluble extract of the plerocercoids were separated using 2-dimensional gel electrophoresis coupled with immunoblot and mass spectrometry analysis. Based on immunoblotting patterns and mass spectrometry results, 8 antigenic proteins were identified from the plerocercoid. Among the proteins, cysteine protease protein might be developed as an antigen for diagnosis of sparganosis. [ABSTRACT FROM AUTHOR]

Abstract Background Sparganosis caused by Spirometra erinaceieuropaei spargana is a zoonotic parasitic infection that has been reported in many countries, including China, Japan, Thailand and Korea, as well as European countries and the USA. The biological and clinical significance of the parasite have previously been reported. Although the genomic and transcriptomic analysis of S. erinaceieuropaei provided insightful views about the development and pathogenesis of this species, little knowledge has been acquired in terms of post-translational regulation that is essential for parasite growth, development and reproduction. Here, we performed site-specific phosphoproteomic profiling, with an aim to obtain primary information about the global phosphorylation status of spargana. Results A total of 3228 phosphopeptides and 3461 phosphorylation sites were identified in 1758 spargana proteins. The annotated phosphoproteins were involved in a variety of biological pathways, including cellular (28%), metabolic (20%) and single-organism (17%) processes. The functional enrichment of phosphopeptides by Gene Ontology analysis indicated that most spargana phosphoproteins were related to the cytoskeleton cellular compartment, signaling molecular function, and a variety of biological processes, including a molecular function regulator, guanyl-nucleotide exchange factor activity, protein kinase activities, and calcium ion binding. The highly enriched pathways of phosphorylation proteins include the phosphatidylinositol signaling system, phagosome, endocytosis, inositol phosphate metabolism, terpenoid backbone biosynthesis, and peroxisome. Domain analysis identified an EF-hand domain and pleckstrin homology domain among the key domains. Conclusions To our knowledge, this study performed the first global phosphoproteomic analysis of S. erinaceieuropaei. The dataset reported herein provides a valuable resource for future studies on the signaling pathways of this important zoonotic parasite.

We report a rare case of sparganosis presented as cauda equina syndrome. Cauda equina syndrome secondary to extramedullary spinal sparganosis infection represents a diagnostic challenge. It should be carefully considered in the differential diagnosis of patients presenting similar characteristics. Although both medical and surgical treatments contribute to the management of this condition, in cases of neurologic deficit due to the mass effect of the lesion, urgent surgical decompression should be attempted to prevent further worsening of the neurological condition.

Spirometra tapeworms (Cestoda: Diphyllobothriidae) collected from carnivorous mammals in Tanzania were identified by the DNA sequence analysis of the mitochondrial cytochrome c oxidase subunit 1 (cox1) and internal transcribed spacer 1 (ITS1), and by morphological characteristics. A total of 15 adult worms were collected from stool samples and carcasses of Panthera leo, Panthera pardus, and Crocuta crocuta in the Serengeti and Selous ecosystems of Tanzania. Three Spirometra species: S. theileri, S. ranarum and S. erinaceieuropaei were identified based on morphological features. Partial cox1 sequences (400 bp) of 10 specimens were revealed. Eight specimens showed 99.5% similarity with Spirometra theileri (MK955901), 1 specimen showed 99.5% similarity with the Korean S. erinaceieuropaei and 1 specimen had 99.5% similarity with Myanmar S. ranarum. Sequence homology estimates for the ITS1 region of S. theileri were 89.8% with S. erinaceieuropaei, 82.5% with S. decipiens, and 78.3% with S. ranarum; and 94.4% homology was observed between S. decipiens and S. ranarum. Phylogenetic analyses were performed with 4 species of Spirometra and 2 species of Dibothriocephalus (=Diphyllobothrium). By both ML and BI methods, cox1 and ITS1 gave well supported, congruent trees topology of S. erinaceieuropaei and S. theileri with S. decipiens and S. ranarum forming a clade. The Dibothriocephalus species were sisters of each other and collectively forming successive outgroups. Our findings confirmed that 3 Spirometra species (S. theileri, S. ranarum, and S. erinaceieuropaei) are distributed in the Serengeti and Selous ecosystems of Tanzania. [ABSTRACT FROM AUTHOR]

Sparganosis is a neglected zoonotic parasitic disease that poses huge threats to humans worldwide. Snakes play an important role in sparganosis transmission because they are the most common second intermediate hosts for Spirometra parasites. However, the population genetics of Spirometra isolates from snakes is currently not well studied in China. The present study was performed to explore the molecular characteristics and phylogenetic analysis of Spirometra tapeworms from different species of snakes in Hunan Province. This study obtained 49 Spirometra isolates from 15 geographical areas in Hunan Province, Central China. Subsequently, the 18S and 28S ribosomal DNA (rDNA) fragments were amplified from the isolated parasites, and their sequences were analyzed to assess their genetic diversity. Phylogenetic analyses were performed using the maximum likelihood algorithm. The results showed that sequence variations among these isolates were 0–2.3% and 0–0.1% for 18S and 28S rDNA, respectively. The phylogenetic analysis showed that all Spirometra isolates from Hunan Province were clustered into the same branch with Spirometra erinaceieuropaei isolated from other areas (China, Vietnam, Australia). Moreover, the phylogenetic trees revealed that Spirometra is closely related to Adenocephalus, Pyramicocephalus, Ligula, Dibothriocephalus, Schistocephalus, and Diphyllobothrium. The Spirometra isolates of different hosts/regions in Hunan Province are not host segregated or geographically isolated, and support for the taxonomic status of Spirometra tapeworms in China has been added. These results provide reference values for future accurate identification and taxonomic status of Spirometra tapeworms in China.

Human sparganosis is a zoonotic disease caused by infection and migration of the plerocercoid of Spirometra spp. Although sparganosis were reported from most parts of the body, the sparganum parasitizing inside cerebral artery is remarkably uncommon. We report a case of cerebral intravascular sparganosis in an elderly patient with acute ischemic stroke who was diagnosed by retrieving sparganum during mechanical thrombectomy. Finally, the parasites were identified as Spirometra erinaceieuropaei using multiplex PCR and cox1 gene sequencing. [ABSTRACT FROM AUTHOR]

Background: Sparganosis caused by Spirometra erinaceieuropaei spargana is a zoonotic parasitic infection that has been reported in many countries, including China, Japan, Thailand and Korea, as well as European countries and the USA. The biological and clinical significance of the parasite have previously been reported. Although the genomic and transcriptomic analysis of S. erinaceieuropaei provided insightful views about the development and pathogenesis of this species, little knowledge has been acquired in terms of post-translational regulation that is essential for parasite growth, development and reproduction. Here, we performed site-specific phosphoproteomic profiling, with an aim to obtain primary information about the global phosphorylation status of spargana. Results: A total of 3228 phosphopeptides and 3461 phosphorylation sites were identified in 1758 spargana proteins. The annotated phosphoproteins were involved in a variety of biological pathways, including cellular (28%), metabolic (20%) and single-organism (17%) processes. The functional enrichment of phosphopeptides by Gene Ontology analysis indicated that most spargana phosphoproteins were related to the cytoskeleton cellular compartment, signaling molecular function, and a variety of biological processes, including a molecular function regulator, guanyl-nucleotide exchange factor activity, protein kinase activities, and calcium ion binding. The highly enriched pathways of phosphorylation proteins include the phosphatidylinositol signaling system, phagosome, endocytosis, inositol phosphate metabolism, terpenoid backbone biosynthesis, and peroxisome. Domain analysis identified an EF-hand domain and pleckstrin homology domain among the key domains. Conclusions: To our knowledge, this study performed the first global phosphoproteomic analysis of S. erinaceieuropaei. The dataset reported herein provides a valuable resource for future studies on the signaling pathways of this important zoonotic parasite. [ABSTRACT FROM AUTHOR]

Mitochondrial DNA sequence variability of Spirometra erinaceieuropaei in GenBank was observed by reinvestigation of mitochondrial cox1 and cytb sequences. The DNA sequences were analyzed in this study, comprising complete DNA sequences of cox1 (n=239) and cytb (n=213) genes. The 10 complete mitochondrial DNA sequences of Spirometra species were compared with those of Korea, China and Japan. The sequences were analyzed for nucleotide composition, conserved sites, variable sites, singleton sites and parsimony-informative sites. Phylogenetic analyses was done using neighbor joining, maximum parsimony, Bayesian inference and maximum-likelihood on cox1 and cytb sequences of Spirometra species. These polymorphic sites identified 148 (cox1) and 83 (cytb) haplotypes within 239 and 213 isolates from 3 Asian countries. Phylogenetic tree topologies were presented high-level confidence values for the 2 major branches of 2 Spirometra species containing S. erinaceieuropaei and S. decipiens, and S. decipiens sub-clades including all sequences registered as S. erinaceieuropaei in cox1 and cytb genes. These results indicated that mitochondrial haplotypes of S. erinaceieuropaei and S. decipiens were found in the 3 Asian countries. [ABSTRACT FROM AUTHOR]

Spargana were collected from human and frogs in Liaoning and Hubei Provinces, China. PCR amplification and direct sequencing of A cox1 fragment was PCR-amplified from genomic DNA extracted from 7 specimens (5 from humans and 2 from frogs). The cox1 fragment (390 bp) showed 97-100% similarity to the reference sequence of S. erinaceieuropaei and 88-89% to the reference sequence of S. decipiens. There were 1-12 bases different between these worms, but no obvious genetic variation (0-3.3%) to the references. There was little difference of cox1 gene between sparganum samples of humans and frogs (1-3%). This study is the first report on S. erinaceieuropaei spargana from humans in Liaoning and Hubei Provinces. [ABSTRACT FROM AUTHOR]

This study was undertaken to determine the complete mitochondrial DNA sequence and structure of the mitochondrial genome of Spirometra ranarum, and to compare it with those of S. erinaceieuropaei and S. decipiens. The aim of this study was to provide information of the species level taxonomy of Spirometra spp. using the mitochondrial genomes of 3 Spirometra tapeworms. The S. ranarum isolate originated from Myanmar. The mitochondrial genome sequence of S. ranarum was compared with that of S. erinaceieuropaei (GenBank no. KJ599680) and S. decipiens (Gen- Bank no. KJ599679). The complete mtDNA sequence of S. ranarum comprised 13,644 bp. The S. ranarum mt genome contained 36 genes comprising 12 protein-coding genes, 22 tRNAs and 2 rRNAs. The mt genome lacked the atp8 gene, as found for other cestodes. All genes in the S. ranarum mitochondrial genome are transcribed in the same direction and arranged in the same relative position with respect to gene loci as found for S. erinaceieuropaei and S. decipiens mt genomes. The overall nucleotide sequence divergence of 12 protein-coding genes between S. ranarum and S. decipiens differed by 1.5%, and 100% sequence similarity was found in the cox2 and nad6 genes, while the DNA sequence divergence of the cox1, nad1, and nad4 genes of S. ranarum and S. decipiens was 2.2%, 2.1%, and 2.6%, respectively. [ABSTRACT FROM AUTHOR]

Copyright of Chinese Journal of Preventive Veterinary Medicine / Zhongguo Yufang Shouyi Xuebao is the property of Chinese Journal of Preventive Veterinary Medicine Editorial Office and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Human sparganosis was diagnosed by morphological and genetic analyses in Korea. The complete mitochondrial genomes of Spirometra erinaceieuropaei and S. decipiens isolated in Korea have been recorded. Present study was performed to provide information to diagnose the etiologic agent of sparganosis by multiplex PCR using mitochondrial genome sequences of S. erinaceieuropaei and S. decipiens. In an effort to examine the differential diagnosis of spirometrid tapeworms, multiplex PCR assays were performed on plerocercoid larvae of S. erinaceieuropaei and S. decipiens. The PCR products obtained using species-specific primers were positively detected in all PCR assays on mixture of S. erinaceieuropaei and S. decipiens DNA. S. erinaceieuropaei-specific bands (239 bp and 401 bp) were obtained from all PCR assays using a mixture of S. erinaceieuropaei-specific primers (Se/Sd-1800F and Se-2018R; Se/Sd-7955F and Se- 8356R) and S. erinaceieuropaei template DNA. S. decipiens-specific bands (540 bp and 644 bp) were also detected in all PCR assays containing mixtures of S. decipiens-specific primers (Se/Sd-1800F and Sd-2317R; Se/Sd-7955F and Sd- 8567R) and S. decipiens template DNA. Sequence analyses on these species-specific bands revealed 100% sequence identity with homologous regions of the mtDNA sequences of S. erinaceieuropaei and S. decipiens. The multiplex PCR assay was useful for differential diagnosis of human sparganosis by detecting different sizes in species-specific bands. [ABSTRACT FROM AUTHOR]

Parasitic helminths modify host immune reactions to promote long-term parasitism. We previously purified a glycoprotein, plerocercoid-immunosuppressive factor (P-ISF), from the excretory/secretory products of Spirometra erinaceieuropaei plerocercoids and reported its cDNA and genomic DNA sequences. In this study, we isolated extracellular vesicles (EVs) from the excretory/secretory products of S. erinaceieuropaei plerocercoids and found that they suppressed the production of nitric oxide and the gene expression of tumor necrosis factor-α, interleukin-1β, and interleukin-6 in lipopolysaccharide-stimulated macrophages. EVs are membrane-bound vesicles 50–250 nm in diameter and are localized in the whole bodies of plerocercoids. EVs from plerocercoids encapsulate a variety of unidentified proteins and microRNAs (miRNAs), which are non-coding RNAs that play essential roles in post-transcriptional gene regulation. The miRNAs of the EVs were analyzed, and 334,137 sequencing reads were mapped to the genomes of other organisms. A total of 26 different miRNA families were identified, including miR-71, miR-10-5p, miR-223, and let-7-5p, which have been reported to have immunosuppressive effects. We confirmed that P-ISF was present in the supernatant but not in the EVs by western blotting with an anti-P-ISF antibody. These results suggest that S. erinaceieuropaei plerocercoids suppress host immunity by releasing P-ISF and EVs. [Display omitted] • Spirometra erinaceieuropaei plerocercoids secreted extracellular vesicles (EVs). • S.erinaceieuropaei EVs suppressed activation of LPS-stimulated macrophages. • EVs contained 26 miRNAs and many unidentified proteins. • EV included the known miRNA (e.g. miR-71) that were reported immuneosuppression. [ABSTRACT FROM AUTHOR]

Although medically important, the systematics of Spirometra and the taxonomic position of S. erinaceieuropaei remain unclear. In this study, the 18S rDNA gene of S. erinaceieuropaei sparganum from naturally infected frogs caught in 14 geographical locations of China was sequenced. In addition, all available 18S sequences of the family Diphyllobothriidae in the Genbank database were included to reconstruct the phylogeny of diphyllobothriid tapeworms. The secondary structure model of the 18S rDNA was also predicated to further explore the sequence variation. Phylogenetic analyses were performed using maximum parsimony (MP), maximum likelihood (ML) and Bayesian inference (BI) methods. The intraspecific divergences of 18S rDNA in Chinese sparganum isolates ranged from 0.0 to 0.4%. Regions of V2, V4 and V7 were the most variable regions in the secondary structure of 18S rDNA. With the exception of genera Duthiersia and Probothriocephalus , other genera (i.e., Adenocephalus , Diphyllobothrium , Diplogonoporus , Duthiersia , Schistocephalus and Spirometra ) selected in the Diphyllobothriidae shared similar topologies of V2, V4 and V7 structures. The topology of generated phylogenetic trees revealed close relationships among Adenocephalus , Digramma , Diphyllobothrium , Diplogonoporus , Ligula , Sparganum and Spirometra . The exact phylogenetic position of Spirometra species should be further analyzed with more sampling and more useful molecular markers. [ABSTRACT FROM AUTHOR]

Sparganosis in humans caused by the plerocercoid larvae of Spirometra erinaceieuropaei is found worldwide, especially in Eastern Asia and the Far East. Previous studies have suggested that dissolution of plerocercoid body, plerocercoid invasion of host tissue, and migration are important processes for sparganosis progression. However, the mechanisms underlying these processes have yet to be determined. Here, we demonstrated the enzymatic property and involvement of a native 23 kDa cysteine protease (Se23kCP), purified from plerocercoids, in sparganosis pathogenesis. Se23kCP is mature protease consisting of 216 amino acids and has a high sequence similarity with cathepsin L in various organisms. Se23kCP conjugated with N -glycans, which have a core fucose residue. Both cysteine and serine protease-specific activities were determined in Se23kCP and their optimal pHs were found to be different, indicating that Se23kCP has a wide range of substrate specificity. Se23kCP was secreted from tegumental vacuoles of the plerocercoid to host subcutaneous tissues and degraded human structural proteins, such as collagen and fibronectin. In addition, the plerocercoid body was lysed by Se23kCP, which facilitated larval invasion of host tissue. Our findings suggest that Se23kCP induces host tissue invasion and migration, and might be an essential molecule for sparganosis onset and progression. [ABSTRACT FROM AUTHOR]

The larval plerocercoid of Spirometra erinaceieuropaei can parasitize humans, causing a serious food borne parasitic zoonosis known as sparganosis. Sparganosis have increased in China in recent years. In this study, the prevalence of sparganum infection in wild frogs in 9 geographical areas in southwest China was firstly investigated. Of 276 caught frogs, 55 frogs were found to be infected with sparganum. Then, the population genetic structure of these sparganum isolates was explored based on four molecular markers ( cytb , cox1 , rrnS and 28S rDNA D1). Highly genetic diversity and the genetic differentiation among sparganum isolates from different sites were revealed in the DNA polymorphism analyses. Both the phylogenetic inference and the analysis of the median-joining network supported two clades in the southwest S. erinaceieuropaei population. However, none demographic population expansion of the southwest S. erinaceieuropaei population was observed in the neutrality test, mismatch distribution analysis and Bayesian skyline plot analysis. Finally, the phylogenetic diversity of S. erinaceieuropaei from eastern, central, southern and southwest China was analyzed, the result suggested that Chinese S. erinaceieuropaei population should be divided into two groups (Group I and Group II), and they started to divergence in the middle Pliocene. [ABSTRACT FROM AUTHOR]

Spirometra erinaceieuropaei is an important diphyllobothriid cestode whose larval plerocercoid (sparganum) can parasitise humans, causing sparganosis. In China, sparganosis is an important foodborne parasitic zoonosis and poses a serious threat to human health. However, our knowledge of the taxonomic classification of S. erinaceieuropaei and its relationship with other diphyllobothriid cestodes, such as Diphyllobothrium , remains fragmentary. In this study, we determined the complete sequences of cytochrome b ( cytb ) and cytochrome c oxidase subunit I ( cox1 ) genes from 61 isolates of S. erinaceieuropaei collected from 11 geographical locations in eastern China and compared them to the sequences of Diphyllobothrium species from the GenBank database. Then, we evaluated the genetic variation of S. erinaceieuropaei and the phylogenetic relationship between S. erinaceieuropaei and Diphyllobothrium . Both negative Tajima’s D and Fu’s F S values of neutrality tests and unimodal curve of mismatch distribution analyses supported demographic population expansion. Bayesian skyline plot (BSP) analyses indicated that a population expansion occurred in the Pleistocene. Phylogenetic analysis based on cytb , cox1 , and cytb + cox1 under all three methods tested (neighbour-joining, maximum parsimony, and maximum likelihood) suggested that the S. erinaceieuropaei isolates from eastern China and the genus Diphyllobothrium are separate monophyletic groups. Furthermore, the uncorrelated log-normal relaxed molecular-clock model suggested that divergence between Spirometra and Diphyllobothrium began in the late Miocene. [ABSTRACT FROM AUTHOR]

The present study was performed to compare the mitochondrial genomes between 2 Spirometra tapeworms, Spirometra erinaceieuropaei and Spirometra decipiens (Cestoidea: Diphyllobothriidae), which larval stages are important etiological agents of sparganosis in humans. For each species, the full mitochondrial genome was amplified in 8 overlapping fragments using total genomic DNA purified from a single worm as the template. The mitochondrial genomes were 13,643 bp (S. erinaceieuropaei) and 13,641 bp (S. decipiens) in length and contained 36 genes; 12 protein-coding genes, 2 ribosomal RNA (rRNA, small and large subunits), and 22 transfer RNAs (tRNAs). The 12 protein-coding genes constituted 10,083 bp (S. erinaceieuropaei) and 10,086 bp (S. decipiens) of their respective mitochondrial genomes. The tRNA genes, ranging in length from 56 to 70 bp, were identified based on putative secondary structures such as the typical cloverleaf shape. A total of 23 intergenic sequences, varying from 1 to 204 bp in size, were interspersed in S. erinaceieuropaei (total, 504 bp) and S. decipiens (total, 496 bp) mtDNA. The 12 protein-coding genes of S. erinaceieuropaei and S. decipiens differed by 12.4%, whereas the overall difference in mtDNA sequence between S. erinaceieuropaei and S. decipiens was 12.9%. Thus, from the standpoint of the mitochondrial genome, S. decipiens represents a valid species that can be distinguished from S. erinaceieuropaei. [ABSTRACT FROM AUTHOR]

Tapeworms of the genus Spirometra are pseudophyllidean cestodes endemic in Korea. At present, it is unclear which Spirometra species are responsible for causing human infections, and little information is available on the epidemiological profiles of Spirometra species infecting humans in Korea. Between 1979 and 2009, a total of 50 spargana from human patients and 2 adult specimens obtained from experimentally infected carnivorous animals were analyzed according to genetic and taxonomic criteria and classified as Spirometra erinaceieuropaei or Spirometra decipiens depending on the morphology. Morphologically, S. erinaceieuropaei and S. decipiens are different in that the spirally coiled uterus in S. erinaceieuropaei has 5-7 complete coils, while in S. decipiens it has only 4.5 coils. In addition, there is a 9.3% (146/1,566) sequence different between S. erinaceieuropaei and S. decipiens in the cox1 gene. Partial cox1 sequences (390 bp) from 35 Korean isolates showed 99.4% (388/390) similarity with the reference sequence of S. erinaceieuropaei from Korea (G1724; GenBank KJ599680) and an additional 15 Korean isolates revealed 99.2% (387/390) similarity with the reference sequences of S. decipiens from Korea (G1657; GenBank KJ599679). Based on morphologic and molecular databases, the estimated population ratio of S. erinaceieuropaei to S. decipiens was 35: 15. Our results indicate that both S. erinaceieuropaei and S. decipiens found in Korea infect humans, with S. erinaceieuropaei being 2 times more prevalent than S. decipiens. This study is the first to report human sparganosis caused by S. decipiens in humans in Korea. [ABSTRACT FROM AUTHOR]

The Spirometra erinaceieuropaei cysteine protease (SeCP) gene encoding a 36 kDa protein was expressed in Escherichia coli, and the potential of recombinant SeCP protein (rSeCP) as an antigen for the serodiagnosis of sparganosis was investigated by ELISA and compared with those of ELISA with sparganum excretory-secretory (ES) antigens. The sensitivity of rSeCP-ELISA and ES-ELISA was 96.67 % (29/30) and 100 % (30/30) respectively, for the detection of anti-sparganum IgG antibodies in sera of the experimentally infected mice ( P > 0.05), and the specificities of both ELISA were 100 % (77/77). In heavily, moderately, and lightly infected mice (five, three, and one larvae per mouse), anti-sparganum antibodies were firstly detected by rSeCP-ELISA at 10-12 days post-infection (dpi), respectively, and then continued to increase with a detection rate of 100 % at 14-22 dpi. In three groups of infected mice, the anti-sparganum antibody levels at different times after infection were statistically different ( P < 0.05). The results showed that the rSeCP might be a potential candidate antigen for early and specific serodiagnosis of sparganosis. But, it needs to be further evaluated with sera of the patients with sparganosis and other helminthiasis. [ABSTRACT FROM AUTHOR]

• A total of 13,659 DEGs were identified in plerocercoid versus adult of S. erinaceieuropaei. • DEGs involved in parasite immunoevasion were more active in plerocercoids. • DEGs associated with metabolic activity were upregulated in adults. The plerocercoid larvae of the tapeworm Spirometra erinaceieuropaei can parasitize humans and animals and cause serious parasitic zoonosis. However, our knowledge of the developmental process of S. erinaceieuropaei is still inadequate. To better characterize differential and specific genes and pathways associated with parasite development, a comparative transcriptomic analysis of the plerocercoid stage and the adult stage was performed using RNA-seq and de novo analysis. Approximately 13,659 differentially expressed genes (DEGs) were identified in plerocercoids versus adults, of which 6455 DEGs were upregulated and 7204 were downregulated. DEGs involved in parasite immunoevasion were more active in plerocercoid larvae than in adults, while DEGs associated with metabolic activity were upregulated in adults. Gene Ontology (GO) and Kyoto Encyclopedia of Genes (KEGG) analyses revealed that most DEGs involved in protein phosphorylation/dephosphorylation and the Wnt signalling pathway were much more active in plerocercoid larvae. The molecular functions of upregulated unigenes in adults were mainly enriched for metabolic activities. qPCR validated that the expression levels of 10 selected DEGs were consistent with those in RNA-seq, confirming the accuracy of the RNA-seq results. Our results contributed to increasing the knowledge on the S. erinaceieuropaei gene repertoire and expression profile and also provide valuable resources for functional studies on the molecular mechanisms of S. erinaceieuropaei. [Display omitted] [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

A 52-year-old woman presented with lower back pain, progressive symmetrical paraparesis with sensory impairment, and sphincter disturbance. Magnetic resonance imaging (MRI) of the whole spine revealed multiple intradural extramedullary serpiginous-mass lesions in the subarachnoid space continuously from the prepontine to the anterior part of the medulla oblongata levels, C7, T2-T8, and T12 vertebral levels distally until the end of the theca sac and filling-in the right S1 neural foramen. Sparganosis was diagnosed by demonstration of the sparganum in histopathological sections of surgically resected tissues and also by the presence of serum IgG antibodies by ELISA. DNA was extracted from unstained tissue sections, and a partial fragment of mitochondrial cytochrome c oxidase subunit 1 (cox1) gene was amplified using a primer set specific for Spirometra spp. coxf. After sequencing of the PCR-amplicon and alignment of the nucleotide sequence data, the causative agent was identified as the larva of Spirometra erinaceieuropaei. [ABSTRACT FROM AUTHOR]