Selective and universal primers for trematode barcoding in freshwater snails

Abstract

Trematodes are significant pathogens of high medical, veterinary, and environmental importance. They are hard to isolate from their intermediate hosts, and their early life stages are difficult to identify morphologically. Therefore, primers were developed for trematodes to create a species barcoding system and allow selective PCR amplification in mixed samples. The specific oligonucleotide primer was universal for trematodes that infected several freshwater snail species in Israel. The diagnostic tool is based on the 18S rDNA gene. In contrast to morphological identification, trematode barcoding is rapid as it is based on a sequence of only 800 bp, and it classifies species accurately due to high polymorphism between conserved areas.

Renal trematode infection in wild birds: histopathological, morphological, and molecular aspects

“…It is also important to consider the role of bird migration in the spread of these parasites. Although there were no reports of migratory birds infected in this study, they have been reported as Paratanaisia hosts (Routtu et al 2014) and could become important carriers, spreading the parasites to locations where bird migration is common.…”

“…The overlap in the morphometric measures of the three species precludes an accurate morphological identification. However, the use of PCR to amplify ribosomal gene fragments and sequencing the products should allow for a proper taxonomic assessment (Routtu et al 2014).…”

“…The GAPDH-positive FFPE samples and parasite specimens were subjected to conventional PCR assays targeting two gene regions, 18S ribosomal DNA (rDNA; Routtu et al 2014) and 28S rDNA (Unwin et al 2013). The first PCR protocol was based on a 310-base pair (bp) fragment of the 28S rDNA gene (Unwin et al 2013).…”

“…The second PCR protocol was based on an 800-bp fragment of the 18S rDNA gene (Routtu et al 2014). This PCR protocol also used 0.25 μL of 0.3-μM forward [C_for (5′-ATGGCTCATTAAATCAGCTAT-3′)] and reverse [A_rev (5′-TGCTTTGAGCACTCAAATTTG-3′)] primers (Síntese Biotecnologia, Belo Horizonte, MG, Brazil) in a reaction mixture otherwise identical to that used for the GAPDH gene.…”

“…This PCR protocol also used 0.25 μL of 0.3-μM forward [C_for (5′-ATGGCTCATTAAATCAGCTAT-3′)] and reverse [A_rev (5′-TGCTTTGAGCACTCAAATTTG-3′)] primers (Síntese Biotecnologia, Belo Horizonte, MG, Brazil) in a reaction mixture otherwise identical to that used for the GAPDH gene. The cycling conditions comprised an initial denaturation of 2 min at 94°C; 30 cycles of 15 s at 94°C, 15 s at 60°C, and 15 s at 72°C; and a final extension of 1 min at 72°C (Routtu et al 2014).…”