Final Report for 2011-2012 Faculty Development Grant
Title: Identification of larval stages of trematodes (Family Opecoelidae) parasitizing freshwater
snails (Family Pleuroceridae)
Abbie M. Tomba
The goal of this project was to identify cryptic larval trematode stages, which parasitize local
freshwater snails, using novel molecular techniques. The three specific objectives of this project
were 1) collect larval and adult trematodes, 2) isolate, amplify and sequence trematode DNA,
and finally 3) analysis the DNA sequences in order to identify larval trematodes, by matching
DNA sequences to adult DNA.
Outcomes and Methodology:
Objective 1 is partially complete. Of the snails collected 5% were parasitized which resulted in
over 20 snail infected with three different types of larval trematodes. Of the 57 fish sampled to
collect adult trematodes, only two fish of a single species (Notropis procne) were found to
contain adult trematodes. While both methods of collection were effective, fish sampling and
processing for parasites is much more time consuming, which resulted a smaller sample size. In
order to get a greater number of adult parasites, fish sampling is ongoing and will be
concentrated on collecting more Notropis procne, which was previously found to be infected.
Objective 2 is partially complete. A good deal of the year was spent designing and optimizing
protocols for DNA isolation and amplification using PCR. The DNA region of interest (ITS
region of rDNA) was successfully isolated and amplified from six larval parasites. Of those six,
reliable sequences were obtained from two samples. Due to the small numbers of adult
trematodes collected, adult trematode DNA has yet to be isolated and amplified.
Work on this
goal is ongoing, with plans to continue isolate and sequence DNA from both larval and adult
trematode samples.
One important aspect of this project was that it gave me the opportunity to learn a suite of
common molecular biology techniques, specifically DNA isolation and PCR. Learning these
techniques was very valuable for me and my students because these methods are being utilized
with increasing frequency in the field of ecology and allow me to answer ecological questions in
new ways.
Objective 3 is partially complete. Because DNA sequences were only obtained from larval
trematodes and not adults, it was not possible to compare sequences between the two different
life stages. However, phylogenetic software (Genius) was used to compare sequences from
obtained larval trematodes to published sequences of other trematodes. As local trematodes are
relatively unstudied, it was not surprising that a close match was not found. However,
phylogenetic analysis supports the initial hypothesis that that one larval specimen was in Family
Opecoelidae. This objective is ongoing and new sequences will be analysed as they are
obtained. Again, another valuable aspect of this project was that I gain experience using
specialized software to compare DNA sequences and produce phylogenetic trees.
Dissemination of Results
My undergraduate research student, Katelynn Monti, also contributed to this project during
summer science 2011 and the 2011-2012 academic year. We presented the preliminary findings
as a poster at the Meeting of the Association of Southeastern Biologists in Athens, GA in April
2012 (abstract and poster attached). I hope to continue with this project over the summer and
upcoming school year, and plan on presenting final results at an upcoming professional meeting.
This grant provided me the opportunities to learn new techniques as well as funds for equipment,
software, supplies and travel, which allowed me to conduct and share this research and will
support future research in my laboratory. I am grateful for the support.