A novel enolase from Taenia solium metacestodes and its evaluation as an immunodiagnostic antigen for porcine cysticercosis.

Cysticercosis is a worldwide parasitic disease of humans and pigs principally caused by infection with the larvae of the pork tapeworm Taenia solium. Through the use of the recently-made-available T. solium genome, we identified a gene within a novel 1448 bp ORF that theoretically encodes for a 433 amino acid-long protein and predicted to be an α-enolase closely related to enolases of other flatworms. Additional bioinformatic analyses revealed a putative plasminogen-binding region on this protein, suggesting a potential role for this protein in pathogenesis. On this basis, we isolated the mRNA encoding for this presumptive enolase from T. solium metacestodes and reverse-transcribed it into cDNA before subsequently cloning and expressing it in both E. coli (rEnoTs) and insect cells (rEnoTsBac), in a 6xHis tagged manner. The molecular weights of these two recombinant proteins were ∼48 and ∼50 kDa, respectively, with the differences likely attributable to differential glycosylation. We used spectrophotometric assays to confirm the enolase nature of rEnoTs as well as to measure its enzymatic activity. The resulting estimates of specific activity (60.000 U/mg) and Km (0.091 mM) are quite similar to the catalytic characteristics of enolases of other flatworms. rEnoTs also exhibited high immunogenicity, eliciting a strong polyclonal antibody response in immunized rabbits. We subsequently employed rEnoTsBac for use in an ELISA aimed at discriminating between healthy pigs and those infected with T. solium. This diagnostic assay exhibited a sensitivity of 88.4% (95% CI, 74.92%-96.11%) and a specificity of 83.7% (95% CI: 69.29%-93.19%). In conclusión, this study reports on and enzymatically characterizes a novel enolase from T. solium metacestode, and shows a potential use as an immunodiagnostic for porcine cysticercosis.

Prevalence of cysticercosis in Estonian pigs and cattle.

Taenia solium has been ranked as the most important foodborne parasite and Taenia saginata as the most commonly found human Taenia tapeworm worldwide. The last official reports of taeniosis from Estonia were in 2003 for T. solium and 2012 for T. saginata. By law, all animal cases of cysticercosis must be registered and reported when found. Our aim was to estimate the prevalence of cysticercosis in Estonia caused by T. solium in pigs and T. saginata in cattle. The four slaughterhouses participating in the study slaughter between them approximately 80% of pigs and cattle in Estonia annually. Sampling spanned from February to April 2014, visiting the slaughterhouses five times per week. Visual inspection, palpation, and incisions at predilection sites were used to find cysts in both species. The sites inspected in both species were the external masseter, tongue, heart, and diaphragm. In addition, the internal masseter in pigs was examined, and the internal pterygoid muscle and esophagus in cattle. DNA was extracted from the cysts and used for PCR amplification of the cox1-gene for Taenia genus and species identification. A total of 564 cattle and 1217 pigs were examined. Cysts were found in 0.36% (n = 2; CI 0.06-1.17) of cattle and in 0.08% (n = 1; CI 0.004-0.40) of pigs. Cestode PCR was negative from all cysts. Results should be considered taking into account the low sensitivity and specificity of finding cysts. Results reflect the situation in larger slaughterhouses, and the possibility that the situation in smaller slaughterhouses is different should not be excluded.

Molecular analyses reveal two geographic and genetic lineages for tapeworms, Taenia solium and Taenia saginata, from Ecuador using mitochondrial DNA.

Tapeworms Taenia solium and Taenia saginata are the causative agents of taeniasis/cysticercosis. These are diseases with high medical and veterinary importance due to their impact on public health and rural economy in tropical countries. The re-emergence of T. solium as a result of human migration, the economic burden affecting livestock industry, and the large variability of symptoms in several human cysticercosis, encourage studies on genetic diversity, and the identification of these parasites with molecular phylogenetic tools. Samples collected from the Ecuadorian provinces: Loja, Guayas, Manabí, Tungurahua (South), and Imbabura, Pichincha (North) from 2000 to 2012 were performed under Maximum Parsimony analyses and haplotype networks using partial sequences of mitochondrial DNA, cytochrome oxidase subunit I (COI) and NADH subunit I (NDI), from Genbank and own sequences of Taenia solium and Taenia saginata from Ecuador. Both species have shown reciprocal monophyly, which confirms its molecular taxonomic identity. The COI and NDI genes results suggest phylogenetic structure for both parasite species from south and north of Ecuador. In T. solium, both genes gene revealed greater geographic structure, whereas in T. saginata, the variability for both genes was low. In conclusion, COI haplotype networks of T. solium suggest two geographical events in the introduction of this species in Ecuador (African and Asian lineages) and occurring sympatric, probably through the most common routes of maritime trade between the XV-XIX centuries. Moreover, the evidence of two NDI geographical lineages in T. solium from the north (province of Imbabura) and the south (province of Loja) of Ecuador derivate from a common Indian ancestor open new approaches for studies on genetic populations and eco-epidemiology.

Identification of species and genetic variation in Taenia isolates from human and swine of North India.

Taenia solium is the major cause of taeniasis and cysticercosis/neurocysticercosis (NCC) in the developing countries including India, but the existence of other Taenia species and genetic variation have not been studied in India. So, we studied the existence of different Taenia species, and sequence variation in Taenia isolates from human (proglottids and cysticerci) and swine (cysticerci) in North India. Amplification of cytochrome c oxidase subunit 1 gene (cox1) was done by polymerase chain reaction (PCR) followed by sequencing and phylogenetic analysis. We identified two species of Taenia i.e. T. solium and Taenia asiatica in our isolates. T. solium isolates showed similarity with Asian genotype and nucleotide variations from 0.25 to 1.01 %, whereas T. asiatica displayed nucleotide variations ranged from 0.25 to 0.5 %. These findings displayed the minimal genetic variations in North Indian isolates of T. solium and T. asiatica.

Human neurocysticercosis case and an endemic focus of Taenia solium in Lao PDR.

A male patient with neurocysticercosis was identified in Montai Village, Xay District, Oudomxay Province, Lao PDR in February 2004. He had a history of diagnosis for neurocysticercosis by a CT scan in Thailand after an onset of epileptic seizure in 1993. A pig in the same district was found to contain Taenia solium metacestodes (=cysticerci); the slaughtered pig body contained more than 2,000 cysticerci. In addition to morphological identification, molecular identification was also performed on the cysticerci by DNA sequencing analysis of the mitochondrial cox1 gene; they were confirmed as T. solium metacestodes. The patient is regarded as an indigenous case of neurocysticercosis infected in an endemic focus of T. solium taeniasis/cysticercosis in Oudomxay Province, Lao PDR.

Characterization of hydrophobic-ligand-binding proteins of Taenia solium that are expressed specifically in the adult stage.

Taenia solium, a causative agent of taeniasis and cysticercosis, has evolved a repertoire of lipid uptake mechanisms. Proteome analysis of T. solium excretory-secretory products (TsESP) identified 10 kDa proteins displaying significant sequence identity with cestode hydrophobic-ligand-binding-proteins (HLBPs). Two distinct 362- and 352-bp-long cDNAs encoding 264- and 258-bp-long open reading frames (87 and 85 amino acid polypeptides) were isolated by mining the T. solium expressed sequence tags and a cDNA library screening (TsHLBP1 and TsHLBP2; 94% sequence identity). They clustered into the same clade with those found in Moniezia expansa and Hymenolepis diminuta. Genomic structure analysis revealed that these genes might have originated from a common ancestor. Both the crude TsESP and bacterially expressed recombinant proteins exhibited binding activity toward 1-anilinonaphthalene-8-sulfonic acid (1,8-ANS), which was competitively inhibited by oleic acid. The proteins also bound to cis-parinaric acid (cPnA) and 16-(9-anthroyloxy) palmitic acid (16-AP), but showed no binding activity against 11-[(5-dimethylaminonaphthalene-1-sulfonyl) amino] undecanoic acid (DAUDA) and dansyl-DL-α-aminocaprylic acid (DACA). Unsaturated fatty acids (FAs) showed greater affinity than saturated FAs. The proteins were specifically expressed in adult worms throughout the strobila. The TsHLBPs might be involved in uptake and/or sequestration of hydrophobic molecules provided by their hosts, thus contributing to host-parasite interface interrelationships.

Morphologic and genetic identification of Taenia tapeworms in Tanzania and DNA genotyping of Taenia solium.

Species identification of Taenia tapeworms was performed using morphologic observations and multiplex PCR and DNA sequencing of the mitochondrial cox1 gene. In 2008 and 2009, a total of 1,057 fecal samples were collected from residents of Kongwa district of Dodoma region, Tanzania, and examined microscopically for helminth eggs and proglottids. Of these, 4 Taenia egg positive cases were identified, and the eggs were subjected to DNA analysis. Several proglottids of Taenia solium were recovered from 1 of the 4 cases. This established that the species were T. solium (n = 1) and T. saginata (n = 3). One further T. solium specimen was found among 128 fecal samples collected from Mbulu district in Arusha, and this had an intact strobila with the scolex. Phylegenetic analysis of the mtDNA cox1 gene sequences of these 5 isolates showed that T. saginata was basal to the T. solium clade. The mitochondrial cox1 gene sequences of 3 of these Tanzanian isolates showed 99% similarity to T. saginata, and the other 2 isolates showed 100% similarity to T. solium. The present study has shown that Taenia tapeworms are endemic in Kongwa district of Tanzania, as well as in a previously identified Mbulu district. Both T. solium isolates were found to have an "African/Latin American" genotype (cox1).

Origin of the pork tapeworm Taenia solium in Bali and Papua, Indonesia.

Global distributions of zoonotic pathogens have been strongly affected by the history of human dispersal and domestication of livestock. The pork tapeworm Taenia solium is distributed worldwide as the cause of neurocysticercosis, one of the most serious neglected tropical diseases. T. solium has been reported in Indonesia but only endemic to restricted areas such as Bali and Papua. Previous studies indicated the distinctiveness of a mitochondrial haplotype confirmed in Papua, but only one isolate has been examined to date. In this study, genetic characterization of T. solium and pigs in Bali and Papua was conducted to clarify the distributional history of the parasite. Mitochondrial haplotype network analysis clearly showed that Indonesian T. solium comprises a unique haplogroup which was the first to diverge among Asian genotypes, indicating its single origin and the fact that it was not introduced in the recent past from other area in Asia in which it is endemic. Although phylogenetic analysis based on the mitochondrial D-loop revealed multiple origins of pigs in Bali and Papua, the majority of pigs belonged to the Pacific Clade, which is widely dispersed throughout the Island Southeast Asia (ISEA) and Oceania due to Neolithic human dispersal. Given the results of our network analysis, it is likely that the Pacific Clade pigs played a key role in the dispersal of T. solium. The data suggest that T. solium was introduced from mainland Asia into Western Indonesia, including Bali, by modern humans in the late Pleistocene, or in the early to middle Holocene along with the Pacific Clade pigs. Introduction into New Guinea most likely occurred in the late Holocene through the spread of Pacific Clade pigs. Over time, T. solium has been eradicated from most of Indonesia through the middle to modern ages owing to religious and cultural practices.

Diagnosis of Taenia solium infections based on "mail order" RNA-sequencing of single tapeworm egg isolates from stool samples.

Combined community health programs aiming at health education, preventive anti-parasitic chemotherapy, and vaccination of pigs have proven their potential to regionally reduce and even eliminate Taenia solium infections that are associated with a high risk of neurological disease through ingestion of T. solium eggs. Yet it remains challenging to target T. solium endemic regions precisely or to make exact diagnoses in individual patients. One major reason is that the widely available stool microscopy may identify Taenia ssp. eggs in stool samples as such, but fails to distinguish between invasive (T. solium) and less invasive Taenia (T. saginata, T. asiatica, and T. hydatigena) species. The identification of Taenia ssp. eggs in routine stool samples often prompts a time-consuming and frequently unsuccessful epidemiologic workup in remote villages far away from a diagnostic laboratory. Here we present "mail order" single egg RNA-sequencing, a new method allowing the identification of the exact Taenia ssp. based on a few eggs found in routine diagnostic stool samples. We provide first T. solium transcriptome data, which show extremely high mitochondrial DNA (mtDNA) transcript counts that can be used for subspecies classification. "Mail order" RNA-sequencing can be administered by health personnel equipped with basic laboratory tools such as a microscope, a Bunsen burner, and access to an international post office for shipment of samples to a next generation sequencing facility. Our suggested workflow combines traditional stool microscopy, RNA-extraction from single Taenia eggs with mitochondrial RNA-sequencing, followed by bioinformatic processing with a basic laptop computer. The workflow could help to better target preventive healthcare measures and improve diagnostic specificity in individual patients based on incidental findings of Taenia ssp. eggs in diagnostic laboratories with limited resources.

The pig tapeworm Taenia solium, the cause of cysticercosis: Biogeographic (temporal and spacial) origins in Madagascar.

Cysticercosis is a serious public health problem in Madagascar. The prevalence rate of active cysticercosis reached 21% in regions with a high level of livestock farming. Taenia solium of African-American and Asian genotypes are both present on the island. The times of divergence of the 13 specimens studied suggests a very ancient diversification of T. solium. These events are widely thought to be prior to the domestication of pigs, and seem to follow the expansion of Homo in Asia. Multiple human migrations and the diversity of potential intermediate hosts may have led to a complex epidemiological situation on the island.

Knowledge, practices and seroprevalence of Taenia species in smallholder farms in Gauteng, South Africa.

Porcine cysticercosis and associated human infections are endemic in Sub-Saharan Africa, Latin America, and Asia. Poor agricultural practices, sanitary practices, and lack of knowledge increase the burden of the diseases in susceptible populations. This study investigates the seroprevalence of Taenia spp. in township pigs in Gauteng, South Africa and describes knowledge and farming practices of pig farmers regarding T. solium infections. Blood samples were collected from 126 pigs in three Gauteng township areas, and analyzed for active Taenia spp. infection using the B158/B60 Ag-ELISA. Farmer questionnaire surveys were conducted in four township areas to investigate the level of knowledge and practices associated with porcine cysticercosis and neurocysticercosis. Logistic regression models were used to assess the relationship between predictor variables and the outcome variable, knowledge of porcine cysticercosis or knowledge of neurocysticercosis. Overall, 7% of the pigs were seropositive for active Taenia spp. infection. 46% of farmers practiced a free-ranging system, while 25% practiced a semi-intensive system. Latrines were absent on all farms; however, 95% of farmers indicated that they have access to latrines at home. Most farmers had no knowledge of porcine cysticercosis (55%) or neurocysticercosis (79%), and this was not associated with any of the factors investigated. The prevalence of active Taenia spp. infection was reasonably low in this study, yet the knowledge level was also low, thus calling for further educational and training programmes to prevent Taenia spp. transmission in these communities.

Taenia solium cysticercosis in Bali, Indonesia: serology and mtDNA analysis.

An active Taenia solium cysticercosis case in Bali, Indonesia, was followed-up by serology and computed tomography. Serology using semi-purified glycoprotein and recombinant antigens showed a drastic drop in titers after calcification of the cysts. Three paraffin-embedded cysts, prepared for histopathological examination, from three other patients were used for mtDNA analysis. The sequences of cox1 gene from T. solium cysticerci from Bali differed from those in Papua and other Asian countries.

Porcine cysticercosis (Taenia solium and Taenia asiatica): mapping occurrence and areas potentially at risk in East and Southeast Asia.

BACKGROUND: Due to the relative short life span and the limited spatial movement, porcine cysticercosis is an excellent indicator of current local active transmission. The aim of this study was to map at province-level, the occurrence of T. solium and T. asiatica in pigs and areas at risk of transmission to pigs in East and Southeast Asia, based on the density of extensive pig production systems and confirmed reports of porcine cysticercosis. METHODS: This study covered East and Southeast Asia, which consist of the following countries: Brunei, Cambodia, China, East Timor, Indonesia, Japan, Laos, Malaysia, Mongolia, Myanmar, North Korea, Philippines, Singapore, South Korea, Thailand and Vietnam. Literature searches were carried out to identify current epidemiological data on the occurrence of porcine cysticercosis caused by T. solium and T. asiatica infections. Modelled densities of pigs in extensive production systems were mapped and compared to available data on porcine cysticercosis. RESULTS: Porcine cysticercosis was confirmed to be present during the period 2000 to 2018 in eight out of the 16 countries included in this study. Taenia solium porcine cysticercosis was confirmed from all eight countries, whereas only one country (Laos) could confirm the presence of T. asiatica porcine cysticercosis. Province-level occurrence was identified in five countries (Cambodia, Indonesia, Laos, Myanmar, and Vietnam) across 19 provinces. Smallholder pig keeping is believed to be widely distributed throughout the region, with greater densities predicted to occur in areas of China, Myanmar, Philippines and Vietnam. CONCLUSIONS: The discrepancies between countries reporting taeniosis and the occurrence of porcine cysticercosis, both for T. solium and T. asiatica, suggests that both parasites are underreported. More epidemiological surveys are needed to determine the societal burden of both parasites. This study highlights a straightforward approach to determine areas at risk of porcine cysticercosis in the absence of prevalence data.

Prevalence and distribution of Taenia solium cysticercosis in naturally infected pigs in Punjab, India.

BACKGROUND: Taenia solium (T. solium) cysticercosis remains a neglected zoonotic disease in India. The current study was planned to estimate the prevalence of T. solium porcine cysticercosis in the Punjab state of India, to compare this prevalence with the disease prevalence in pigs reared outside Punjab and to assess the distribution of the parasite in pig carcasses. METHODS: Two slaughter shops were selected in each of the 22 districts of Punjab. Pigs slaughtered on the day/s of inspection were post-mortem inspected to identify the presence of T. solium cysts. Estimated true prevalence was estimated by taking into account the diagnostic sensitivity (38%) and specificity (100%) of post-mortem inspection using the Rogan-Gladen estimator. Positive carcasses were purchased and brought to the laboratory to assess the tissue distribution of T. solium cysts and to conduct PCR targeting large subunit rRNA gene, internal transcribed spacer 1 gene, ITS1 gene and Cytochrome oxidase I gene. The selected PCR products were submitted for sequencing and phylogenetic analyses were performed. FINDINGS: We contacted 71 shop owners to achieve a sample of 44 shops for the study. We inspected 642 pigs reared in Punjab and 450 imported from other states at these slaughter shops. In addition, we sampled 40 pigs from an abattoir located in the state capital. Of the 642 pigs reared in Punjab, 9 had T. solium cysts with an apparent prevalence of 1·40% (95% CI: 0·74%, 2·64%) and the estimated true prevalence of 3.69% (95% CI: 1·95%, 6·95%). Pigs imported from outside the state had a significantly higher prevalence (odds ratio: 2·58; 95% CI: 1·12, 5·98; p-value: 0·026) as 15 of the 450 imported pigs were positive (apparent prevalence: 3.33%; 95% CI: 2.03%, 5.43%; estimated true prevalence: 8.77%; 95% CI: 5.34%, 14.28%). None of samples was positive from the pigs sampled at the abattoir in the state capital. The PCR confirmed T. solium cysts from all the 24 positive samples. We counted a median of 897 (range 526-1964) cysts per infected pig from the 19 infected pig carcasses inspected. The phylogenetic tree based on the alignment of partial cytochrome oxidase 1 sequences indicated all positive samples to be clustered with the T. solium Asian genotype. The analysis did not indicate the presence of T. asiatica in the slaughter pigs. CONCLUSIONS: Despite the underestimation of the prevalence due to missing mildly-infected carcasses, low participation and lack of representative sampling, the presence of heavily infected carcasses containing viable cysts, particularly those imported from outside the state, indicates that T. solium cysticercosis is an important food safety concern for pork consumers in Punjab, India. Measures should be taken to reduce the disease prevalence in pigs to reduce the disease burden in the public.

Complete sequence of the mitochondrial genome of Taenia saginata: comparison with T. solium and T. asiatica.

The complete sequence of the Taenia saginata mitochondrial genome was determined, and its organization and structure were compared to other human-tropic Taenia tapeworms for which complete mitochondrial sequence data were available. The mitochondrial genome was 13,670 bp long, contained 12 protein-coding genes, two ribosomal RNAs (rRNAs, a small and a large subunit), and 22 transfer RNAs (tRNAs). It did not encode the atp8 gene. Overlapping regions were found between nad4L and nad4, nad1 and trnN, and cox1 and trnT. The ATG initiation codon was used for 10 protein-coding genes, and the GTG initiation codon was used for the remaining 2 genes (nad4 and atp6). The size of the protein-coding genes of the three human Taenia tapeworms did not vary, except for Taenia solium nad1 (891 aa) and nad4 (1212 aa) and Taenia asiatica cox2 (576 aa). The tRNA genes were 57-75 bp long, and the predicted secondary structures of 18 of these genes had typical clover-leaf shapes with paired dihydrouridine (DHU) arms. The genes in all human Taenia tapeworms for the two mitochondrial rRNA subunits rrnL and rrnS are separated by trnC. The putative T. saginata rrnL and rrnS are 972 and 732 bp long, respectively. The non-coding regions of the mt genome of T. saginata consisted of 2 regions: a short non-coding region (SNR, 66 nucleotides) and a long non-coding region (LNR, 159 nucleotides). The overall sequence difference in the full mitochondrial genome between T. saginata and T. asiatica was 4.6%, while T. solium differed by 11%. In conclusion, the complete sequence of the T. saginata mitochondrial genome will serve as a resource for comparative mitochondrial genomics and systematic studies of the parasitic cestodes.

Fasciclin-calcareous corpuscle binary complex mediated protein-protein interactions in Taenia solium metacestode.

BACKGROUND: Neurocysticercosis (NC) caused by Taenia solium metacestode (TsM) is a serious neurological disease of global concern. Diverse bioactive molecules involved in the long-term survival of TsM might contribute to disease progression. Fasciclin (Fas) is an extracellular protein that mediates adhesion, migration and differentiation of cells by interacting with other molecules. We hypothesized that TsMFas might bind to calcareous corpuscle (CC) through its adhesive property and participate in crucial protein-protein interactions, thus contributing to the creation of a symbiotic interactome network. METHODS: Two paralogous TsMFas (TsMFas1 and TsMFas2) were isolated, and their molecular properties were characterized. The co-localization pattern of TsMFas1 and TsMFas2 with CC was determined. CC-TsMFas binary complex was generated by incubating CC with recombinant proteins (rTsMFas1 and 2). In vitro binding assay of CC-rTsMFas1 or CC-rTsMFas2 binary complex with TsM cellular proteins extracted from scolex and neck was conducted. Their binding partners were identified through proteomic analysis. Integrated protein-protein interaction networks were established. RESULTS: TsMFas1 (6072 bp long) was composed of 15 exons (841 amino acid polypeptide) interrupted by 14 introns. TsMFas2 (5201 bp long) comprised of 11 exons (597 amino acids) and 10 intervening introns. These proteins displayed 22% amino acid sequence identity to each other, but tightly conserved Fas-related domains. Several isoforms of Fas1 and Fas2 proteins might have been expressed through post-translational modifications. They showed adhesion activity with other cells. TsMFas proteins were largely distributed in parenchymal regions of the scolex and bladder wall. These molecules were co-localized with CC, a unique organelle found in platyhelminths. Subsequent proteome analysis of CC-Fas binary complex mediated protein-protein interactions revealed seven protein ligands in the TsM cellular proteins. Their functions were mainly segregated into carbohydrate metabolism (enolase, phosphoenolpyruvate carboxykinase, phosphoglycerate kinase and glyceraldehyde 3-phosphate dehydrogenase) and cytoskeleton/cellular motility (actin, paramyosin and innexin nuc-9). Those proteins had direct (physical) and/or indirect (functional) relationships along with their biochemical properties and biological roles. CONCLUSION: Protein repertoires strongly suggest that TsMFas and CC may symbiotically mediate protein-protein interactions during biological processes to maintain efficacious homeostatic functions and ensure the prolonged survival of TsM in the host.

Neurocysticercosis: A case study of a Mongolian traveler who visited China and India with an updated review in Asia.

BACKGROUND: Neurocysticercosis (NCC) of Taenia solium has never been reported in Mongolia. A Mongolian traveler who visited China and India presented with epileptic seizures after his return to Mongolia. Magnetic resonance imaging showed multiple ring enhancing lesions with edema surrounding several lesions in his brain that were initially diagnosed to be viral encephalitis or NCC. METHODS: Serology for cysticercosis using diagnostic antigens partially purified by cation-exchange chromatography and chimeric recombinant antigens and DNA analysis of a proglottid found in the patient's stool were applied for differential diagnosis. RESULTS: Serology showed strong positivity for NCC, and mitochondrial cox1 gene analysis revealed Indian but not Chinese haplotype of T. solium. CONCLUSION: This NCC case was considered to be caused by eggs released from adult tapeworm(s) established after eating uncooked or undercooked pork contaminated with cysticerci during his stay in India. This is a case report of taeniasis and secondary NCC associated with travel to India with an updated review of NCC in Asia. Molecular tracking of the specimen is highly informative as a way to identify where the infection was acquired.

Genetic variation in Taenia solium.

Neurocysticercosis is a major zoonotic larval cestode infection that has a worldwide distribution and is of significant public health importance. Knowledge of the genetic structure of Taenia solium can be applied to the epidemiology and transmission of this disease, since genetic variants may differ in infectivity and pathogenicity. Molecular epidemiological approaches can also enable detailed studies of transmission. On a global scale, mitochondrial markers have differentiated between T. solium isolates from Asia and isolates from Africa/Latin America. Intraspecific variation in T. solium has been detected to some extent, using RAPD markers to differentiate between T. solium populations from different regions within Mexico. Markers currently available for T. solium have not been used to analyse genetic variation at the community or local level. The development of highly polymorphic markers such as microsatellites in T. solium can provide the means to examine genetic heterogeneity of tapeworm infection at the household, community and regional level. Preliminary studies suggest it is possible to analyse population genetic variation in communities using a range of polymorphic markers.

Fecal specimens preparation methods for PCR diagnosis of human taeniosis.

Sample preparation and DNA extraction protocols for DNA amplification by PCR, which can be applied in human fecal samples for taeniasis diagnosis, are described. DNA extracted from fecal specimens with phenol/chloroform/isoamilic alcohol and DNAzol reagent had to be first purified to generate fragments of 170 pb and 600 pb by HDP2-PCR. This purification step was not necessary with the use of QIAmp DNA stool mini kit. Best DNA extraction results were achieved after eggs disruption with glass beads, either with phenol/chloroform/isoamilic alcohol, DNAzol reagent or QIAmp DNA stool mini kit.

Recent advances and perspectives in molecular epidemiology of Taenia solium cysticercosis.

Cysticercosis caused by accidental ingestion of eggs of Taenia solium is spreading all over the world through globalization and is one of the most neglected, neglected tropical diseases (NTDs) or neglected zoonotic diseases (NZDs). In the present study, the reason why T. solium cysticercosis has been neglected is discussed at first, and followed with an overview on the most recent advances and perspectives in molecular approaches for epidemiology of T. solium taeniasis/cysticercosis, since although taeniasis does not constitute recognized zoonoses, transmission and complete development are dependent on human definitive hosts. Main topics are discussions on (1) the two, Asian and Afro/American, genotypes of T. solium, (2) comparative analysis of mitochondrial (haploid) and nuclear (diploid) genes, and (3) the presence of hybrids of these two genotypes which indicates out-crossing of two genotypes in hermaphrodite tapeworms in Madagascar. Additional topics are on (4) the usefulness of phylogeographic analyses to discuss where the infection was acquired from, and (5) miscellaneous unsolved topics around these genetic diversity of T. solium.