ITS-2 rDNA SEQUENCING OF<i>GNATHOSTOMA</i>SPECIES (NEMATODA) AND ELUCIDATION OF THE SPECIES CAUSING HUMAN GNATHOSTOMIASIS IN THE AMERICAS

Almeyda-Artigas, Roberto Javier; Bargues, M. D.; Mas‐Coma, Santiago

doi:10.1645/0022-3395(2000)086[0537:irsogs]2.0.co;2

Cited by 119 publications

(23 citation statements)

References 31 publications

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“…Primers, NEWS2 (forward) 5′-TGTGTCGATGAAGAACGCAG-3′ and ITS2-RIXO (reverse) 5′-TTCTATGCTTAAATTCAGGGG-3′ were used to amplify a 600-bp fragment of the 5.8S rRNA gene and the intergenic transcribed spacer 2 (ITS-2) by using PCR ( 30 ) to corroborate morphologic identifications. Five microliters of the reaction mixture was examined by 1% agarose gel containing 0.0001% Gel Red (Phenix Research Products, Candler, NC, USA) by gel electrophoresis.…”

Section: Methodsmentioning

confidence: 99%

“…These sequences were aligned along with 20 sequences of Gnathostoma spp. available on GenBank: G. spinigerum , G. binucleatum , G. hispidum , G. nipponicum , G. miyazakii , G. lamothei , G. doloresi , and G. turgidum from NCBI Database using ClustalW version 5.1 in MEGA ( 30 ) and manually trimmed to remove overhang. Sequences from this study were deposited in GenBank under accession nos.…”

Section: Methodsmentioning

confidence: 99%

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Gnathostoma spinigerumin Live Asian Swamp Eels (Monopterusspp.) from Food Markets and Wild Populations, United States

Cole¹,

Choudhury²,

Nico³

et al. 2014

Emerg. Infect. Dis.

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In Southeast Asia, swamp eels (Synbranchidae: Monopterus spp.) are a common source of human gnathostomiasis, a foodborne zoonosis caused by advanced third-stage larvae (AL3) of Gnathostoma spp. nematodes. Live Asian swamp eels are imported to US ethnic food markets, and wild populations exist in several states. To determine whether these eels are infected, we examined 47 eels from markets and 67 wild-caught specimens. Nematodes were identified by morphologic features and ribosomal intergenic transcribed spacer–2 gene sequencing. Thirteen (27.7%) M. cuchia eels from markets were infected with 36 live G. spinigerum AL3: 21 (58.3%) in liver; 7 (19.4%) in muscle; 5 (13.8%) in gastrointestinal tract, and 3 (8.3%) in kidneys. Three (4.5%) wild-caught M. albus eels were infected with 5 G. turgidum AL3 in muscle, and 1 G. lamothei AL3 was found in a kidney (both North American spp.). Imported live eels are a potential source of human gnathostomiasis in the United States.

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Gnathostoma spinigerumin Live Asian Swamp Eels (Monopterusspp.) from Food Markets and Wild Populations, United States

Cole¹,

Choudhury²,

Nico³

et al. 2014

Emerg. Infect. Dis.

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“…The primers used were News2 5′-TGT-GTC-GAT-GAA-GAA-CGC-AG-3′ and Its2Rixo 5′-TTC-TAT-GCT-TAA-ATT-CAG-GGG-3′ [3, 6]. The sequences were amplified using a commercial kit (Taq PCR Master Mix, Qiagen) in a total volume of 25 μL (containing 3 mM of MgCl 2 and 400 μM of each dNTP) in a Peltier Thermal Cycler (MJ Research) with an initial denaturation step at 94 °C for 3 min, followed by 40 cycles, each comprising denaturation at 94 °C for 30 s, annealing at 55 °C for 30 s, extension at 72 °C for 45 s, and a final extension step at 72 °C for 5 min.…”

Section: Methodsmentioning

confidence: 99%

“…The primers used for amplification of Fasciola sp. sequences were Fsh1 5′-GAT-CAA-TTC-ACC-CAT-TTC-CGT-TAG-TCC-TAC-3′ and Fsh2 5′-AAA-CTG-GGC-TTA-AAC-GGC-GTC-CTA-CGG-GCA-3′ and for lymnaeids ITS - 2 amplification sequences were News2 5′-TGT-GTC-GAT-GAA-GAA-CGC-AG-3′ and Its2Rixo 5′-TTC-TAT-GCT-TAA-ATT-CAG-GGG-3′ [3, 6]. The sequences were amplified using a commercial kit (Taq PCR Master Mix, Qiagen) in a total volume of 25 μL in a Peltier Thermal Cycler (MJ Research) with an initial denaturation step at 95 °C for 5 min, followed by 40 cycles, each comprising denaturation at 95 °C for 1 min, annealing at 56 °C for 1 min, extension at 72 °C for 1 min, and a final extension step at 72 °C for 10 min.…”

Section: Methodsmentioning

confidence: 99%

IsGalba schirazensis(Mollusca, Gastropoda) an intermediate host ofFasciola hepatica(Trematoda, Digenea) in Ecuador?

Caron¹,

Celi-Erazo²,

Hurtrez‐Boussès³

et al. 2017

Parasite

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Fasciolosis is a widely distributed disease in livestock in South America but knowledge about the epidemiology and the intermediate hosts is relatively scarce in Ecuador. For three months, lymnaeid snails were sampled (n = 1482) in Pichincha Province at two sites located in a highly endemic area. Snails were identified (based on morphology and ITS-2 sequences) and the infection status was established through microscopic dissection and a multiplex polymerase chain reaction (PCR)-based technique. Techniques based on morphology were not useful to accurately name the collected snail species. Comparison with available DNA sequences showed that a single snail species was collected, Galba schirazensis. Live rediae were observed in 1.75% (26/1482) and Fasciola sp. DNA was detected in 6% (89/1482) of collected snails. The COX-1 region permitted identification of the parasite as Fasciola hepatica. The relative sensitivity and specificity of the microscope study, compared to PCR results, were 25.84% and 99.78%, respectively. The mean size of the snails recorded positive for F. hepatica through crushing and microscopy was significantly higher than the mean size of negative snails, but there was no such difference in PCR-positive snails. The role of G. schirazensis as an intermediate host of F. hepatica in Ecuador is discussed and the hypothesis of an adaptation of the parasite to this invasive snail is proposed. For the first time, an epidemiological survey based on molecular biology-based techniques assessed the possible role of lymnaeid snails in the epidemiology of fasciolosis in Ecuador.

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“…DNA concentration was measured by spectrophotometer ND-1000 (NanoDrop®) and then the samples were stored at -20°C. PCR was carried out in total volume of 25 μl using PPP Master Mix (Top-Bio), 50 ng of snail template DNA and 1 μl of 10 μM forward primer 5′ - TGT GTC GAT GAA GAA CGC AG – 3′ and reverse primer 5′ - TTC TAT GCT TAA ATT CAG GGG - 3′ specific for 502 bp ITS2 region [32]. …”

Section: Methodsmentioning

confidence: 99%

Highland cattle and Radix labiata, the hosts of Fascioloides magna

et al. 2014

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BackgroundFascioloides magna is a pathogenic fluke introduced to Europe ca 140 years ago. As it is spreading over the continent, new intermediate and definitive hosts might be involved in transmission of the parasite. In Europe, several studies reported potential new intermediate snail hosts (Radix spp.) for F. magna, and also several cases of fascioloidosis of wild and domestic animals were published. However, the data based on molecular and histological analyses confirming these findings remained unreported. This study aims to refer to unique findings of F. magna in European snails and domestic animals (the first observation in the Czech Republic in the last 30 years) and demonstrate the use of molecular techniques in determination of F. magna.ResultsTwo snails of R. labiata naturally infected with F. magna were found; mature cercariae and daughter rediae were observed. Maturity of cercariae was checked by histological methods, however, their ability to encyst was not confirmed. Co-infection of F. magna and Fasciola hepatica in the liver of two highland cattle bulls was proved. Adult fasciolid flukes producing eggs were found in the liver pseudocysts (F. magna) and the bile ducts (F. hepatica). Identification of intermediate hosts, intramolluscan stages, adult flukes and eggs was performed by sequencing the ITS2 region. Connection of F. magna pseudocysts with the gut (via the bile ducts) was not confirmed by means of histological and coprological examinations.ConclusionsFor the first time, Radix labiata was confirmed as the snail host for F. magna under natural conditions and, together with the finding of F. magna infection in cattle, we can expect further transmission of F. magna from wildlife to livestock in localities shared by these hosts.

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ITS-2 rDNA SEQUENCING OFGNATHOSTOMASPECIES (NEMATODA) AND ELUCIDATION OF THE SPECIES CAUSING HUMAN GNATHOSTOMIASIS IN THE AMERICAS

Cited by 119 publications

References 31 publications

Gnathostoma spinigerumin Live Asian Swamp Eels (Monopterusspp.) from Food Markets and Wild Populations, United States

Gnathostoma spinigerumin Live Asian Swamp Eels (Monopterusspp.) from Food Markets and Wild Populations, United States

IsGalba schirazensis(Mollusca, Gastropoda) an intermediate host ofFasciola hepatica(Trematoda, Digenea) in Ecuador?

Highland cattle and Radix labiata, the hosts of Fascioloides magna

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