Analysis of Cliff Swallow’s Diet via DNA Barcoding and Bioinformatics James Corbitt, Rebecca Bednorz, and Joanna Werner-Fraczek Moreno Valley College, CA

Abstract Materials & Method Results Discussion Moreno Valley College (MVC) is a seasonal home to During the 2016 season, four Cliff Swallows were identified by DNA Barcoding The presented data represents a narrow view on migratory birds, Cliff Swallows (Petrochelidon found dead. Through dissection and light microscopy, Taxonomic name Common name swallows’ diet since the dead birds were found pyrrhonota), that build their nests on campus the buccal cavity, esophagus, and stomach were Empoasca vitis Leafhopper within two weeks. This does not indicate the buildings. Cliff swallows are insectivores that inspected for any remains. For a thorough Sciaridae Darked winged fungus gnats possible season fluctuation in the diet because of consume thousands of insects a day. The analysis of analysis, the stomach was removed and all contents Sciaridae Darked winged fungus gnats different insects being available at different times. the swallow's diet considering their predatory nature were examined under the microscope. To further Mycetophilidae Fungus gnats More studies are needed. will help to assess the role of swallows as natural separate and clean the different pieces of insects for Agrotis subterranea Granulate cutworm regulators of the insect population. This study report a more accurate DNA sequence, the pieces for DNA Harpalus honestus Ground beetle In order to move away from depending on dead on insect species found in the stomach of swallows, isolation were placed on a paper towel then filtered sericata Common green bottle birds, we are in the process of developing a flying insects present on the MVC campus, as well using distilled water. Insect remains that observed to Lucilia sericata Common green bottle fly procedure where bird's droppings will be analyzed. as ecoparasites found in the nests. DNA barcoding be suitable were extracted for DNA isolation. Lucilia sericata Common green bottle fly Droppings would be collected throughout the was the selected method to identify insects via Following DNA isolation, the samples were gel Lucilia sericata Common green bottle fly season in one week intervals at 2-3 locations. DNA bioinformatics tools. Twenty-four insect species were purified and DNA barcoded (i.e. the “648 base-pair Lucilia sericata Common green bottle fly will be isolated and amplified with genus/species identified from the insects captured on the MVC region in the mitochondrial cytochrome c oxidase 1 Lucilia sericata Common green bottle fly specific primers. This would provide information if campus. Eight insect species were found in the gene ‘CO1’” gets amplified using Folmer Primers for Lucilia sericata Common green bottle fly the insect in question, known to populate Moreno collected nests. Future quantitative studies will PCR) (Barcode of Life 2017; Butler et al. 2012). The Lucilia sericata Common green bottle fly Valley area, is consumed by cliff swallows. address the correlation between the presence of results were sent to Eton Bioscience Inc. (San Diego Lucilia sericata Common green bottle fly swallows and the insect population at MVC. based biotech company) for DNA sequencing. Lucilia sericata Common green bottle fly Subsequently, FinchTV (bioinformatics software by Australian blowfly References Figure 1: (Left) The Geospiza) was used to read chromatogram files and Lucilia cuprina Australian sheep blowfly Barcode of Life. 2017. “What is DNA Barcoding?” n.d. Swallow was located during the clean the sequences, then inputted into the NCBI Lucilia cuprina Australian sheep blowfly www.barcodeoflife.org/content/about/what-dna- 2016 spring season BLAST database which compares nucleotide Lucilia cuprina Australian sheep blowfly barcoding. and was dissected sequences and computes statistical similarities. This chart represents 25% of the samples that were on June 23rd, 2016. Butler, Mandy. Heather Henter, and Stephanie Mel. By means of taken. The remaining were not included due to 2012. From Bugs to Barcodes: Using Molecular Tools to dissection, the contamination. As a result, a different approach is buccal cavity, Study Biodiversity. Division of Biological Sciences, esophagus, and being taken that involves using specific genus / UCSD. sdbiodiversity.ucsd.edu/info/index.html stomach were Figure 2: (Left) species primers. inspected for any This Cliff Figure 4: (Right) The DNA Sequencing Services. Eton Bioscience insects. For further Swallow’s picture on the right shows study, the stomach stomach was incorporated. www.etonbio.com/. the result of separating was completely extracted on May charged molecules (in this removed for all 13th, 2016. The case DNA) through the Millham, Cheryl. 2012. “Swallows may be messy, but contents to be contents were technique called gel they keep the mosquitoes away.” examined. Results separated using electrophoresis. The were separated for light microscopy www.laketahoenews.net/2012/05/swallows-may-be- combination of a loading DNA isolation. and dissecting messy-but-they-keep-the-mosquitoes-away/. dye and the ultra-violet utensils. Introduction lighting causes the DNA to As springtime approaches, Cliff Swallows migrate be more visible and easier National Center for Biotechnology Information. to extract as the DNA Standard Nucleotide BLAST. U.S. National Library of from the surrounding areas of Central and South seems fluorescent. Each Medicine. n.d. America to breed. Resultantly, many of these birds agarose gel contains eight samples of insect DNA blast.ncbi.nlm.nih.gov/Blast.cgi?PAGE_TYPE=BlastSea reside in Southern California – particularly Moreno with a total of 48 samples. This was completed on October 3rd, 2016. rch. Valley – for the duration of their stay. Since Southern California has been growing in agriculture, industry, Acknowledgements and residential living, these birds have become an Figure 3: (Right) The research was supported by CCURI that is essential subject of study. Swallows are secondary to The image shows funded through a National Science Foundation tertiary consumers in the local ecosystem as the separated grant (#1118679), and by MVC independent insectivores which consume numerous insects per pieces of insects retrieved from the NSF grant (#1525847). day. Due to this intrinsic nature, these birds can stomach of the serve as biological controls of certain insect Cliff Swallow. The populations. One study cites that Cliff Swallows “can insect located at the center of the eat up to 1,000 mosquitoes in a day — per bird” picture appears to (Millham, 2012). Consequently, this fluctuation can be the Western be beneficial or harmful to agriculture, humans, and drywood termite (Incisitermes related ecology in the surrounding area. This specific minor). research being conducted at Moreno Valley College Figure 5: (Above) The following picture depicts a Lucilia sericata result is to analyze the diet of Cliff Swallows in order to that is being examined with FinchTV. Specific portions of the DNA determine the specific insects being consumed. sequence can be selected and ran through the NCBI BLAST database.