A Ribeiroia spp. (class: Trematoda)--specific PCR-based diagnostic.

Area of Science:

• Parasitology
• Molecular Biology
• Ecology

Background:

• Amphibian populations are declining globally, with increased reports of malformations in North America.
• Ribeiroia ondatrae is a known cause of amphibian malformations, but its prevalence in North America is poorly understood.

Purpose of the Study:

• To develop a rapid, accurate, and cost-effective diagnostic tool for detecting Ribeiroia spp. infection in snails.
• To facilitate field studies on the prevalence and impact of Ribeiroia spp. on amphibian populations.

Main Methods:

• Designed oligonucleotide primers targeting unique sequences in the ribosomal internal transcribed spacer-2 region of Ribeiroia spp.
• Developed a single-tube polymerase chain reaction (PCR) assay for detecting parasite DNA in Planorbella trivolvis snails.
• Optimized PCR sensitivity to detect DNA from a single R. ondatrae cercaria and developed a nested PCR for enhanced sensitivity.

Main Results:

• The PCR assay reliably produced a 290-bp product, detecting less than one cercariae equivalent.
• Nested PCR detected as little as 100 fg of the target DNA fragment.
• The assay showed specificity, not detecting DNA from other trematode species.
• Field testing of 137 snails identified 3 positive cases, with results consistent with microscopic examination.

Conclusions:

• The developed PCR diagnostic is a sensitive, specific, and cost-effective tool for identifying Ribeiroia spp. infections in snails.
• This diagnostic will significantly aid in monitoring parasite prevalence and understanding its role in amphibian malformations.
• The method offers a valuable tool for ecological studies investigating host-parasite dynamics in amphibian populations.