A Comparison of Standard Clinical Microbiology Diagnostic Techniques to High Throughput Sequencing Using 16s rRNA Gene Analysis in Wild, Sea Side and Aquarium Bottlenose Dolphins (Tursiops truncatus), and the Water They Inhabit
Abstract
The relevance of bacterial population involvement in health and disease of animals and humans has been relatively recent following the early development of microbiology in the mid-19th century. The information gained from clinical cultures is often incomplete due to cost and the resources needed, limits set on species tested, drugs available in sensitivity panels, and availability to detect anaerobic organisms. The implications of the relationship between emerging microbiota research and health including immune system support, digestion, protection from pathogenic bacteria,2,3,4,5 and an appropriate balanced bacterial flora6 in each body site is thought to be extremely important. To better document and compare bacterial populations of interest in bottlenose dolphins, samples were collected from skin, blow, gastric, fecal and urogenital sites from dolphins from an aquarium, seaside pens, and Sarasota Bay during health examinations.1 These were analyzed by standard cultures techniques at Micrim Labs Inc. and were compared to parallel samples assessed by PCR amplification of a hypervariable region (V3–V4) of the bacterial 16s rRNA gene for each sample followed by sequencing on an Illumina MiSeq platform.2 Water samples were also obtained from each location for microbiota. Results showed a surprising variation between populations and environments compared to standard culture techniques and the need to consider anaerobes as a major component of normal flora and therapy. Veterinarians should consider the technique’s potential for future inclusion in clinical applications as normal ranges are determined, prices go down and this area better explains numerous aspects of the animal’s bacterial populations.
Acknowledgments
The authors would like to express their deep gratitude to the researchers, water quality personnel, veterinarians and staff members at the National Aquarium, Dolphin Research Center, Clearwater Marie Aquarium and the Sarasota Dolphin Research Program for their dedication and support for this project. This project was financially supported by the John Ben Snow Foundation, Syracuse NY.
Micrim Labs Inc. 800 E Cypress Creek Rd # 202, Fort Lauderdale, FL 33334
*Presenting author
Literature Cited
1. Wells RS, Rhinehart HL, Hansen LJ, Sweeney JC, Townsend FI, Stone R, Casper D, Scott MD, Hohn AA, Rowles TK. 2004. Bottlenose dolphins as marine ecosystem sentinels: Developing a health monitoring system. EcoHealth 1:246–254.
2. Bik EM, Costello EK, Switzer, AD, Callahan BJ, Holmes SP, Wells RS, Carlin KP, Jensen ED, Venn-Watson S, Relman DA. 2016. Marine mammals harbor unique microbiotas shaped by and yet distinct from the sea. Nature Commun. 7:10516 doi: 10.1038/ncomms 10516.
3. Hanning I, Diaz-Sanchez S. 2015. The functionality of the gastrointestinal microbiome in non-human animals. Microbiome. 3:51 DOI 10.1186/s40168-015-0113.
4. Shulzhenko N, Morgun A, Hsiao W, Battle M, Yao M, Gavrilova O, Orandle M, Mayer L, Macpherson AJ, McCoy KD, Fraser-Liggett C, Matzinger P. 2011. Crosstalk between B lymphocytes, microbiota and the intestinal epithelium governs immunity versus metabolism in the gut. Nature Medicine, 17(12), 1585–93. doi:10.1038/nm.2505.
5. Tlaskalova-Hogenova H, Stepankova R, Hudcovic T, Tuckova L, Cukrowska B, Lodinova-Zadnikova R, Kozakova H, Rossmann P, Bartova J, Sokol D, Funda D.P, Borovska D, Rehakova Z, Sinkora J, Hofman J, Drastich P, Kokesova A. 2004. Commensal bacteria (normal microflora), mucosal immunity and chronic inflammatory and autoimmune diseases. Immunol Lett. 93:97–108.
6. Diaz MA, Bik EM, Carlin KP, Venn-Watson SK, Jensen ED, Jones SE, Gaston EP, Relman DA, Versalovic J. 2013. Identification of Lactobacillus strains with probiotic features from the bottlenose dolphin (Tursiops truncatus). J. Appl. Microbiol. 115:1037–1051.