Date Published: April 22, 2019
Publisher: Public Library of Science
Author(s): Martha Valdez-Moreno, Natalia V. Ivanova, Manuel Elías-Gutiérrez, Stephanie L. Pedersen, Kyrylo Bessonov, Paul D. N. Hebert, Hideyuki Doi.
Environmental DNA (eDNA) is an effective approach for detecting vertebrates and plants, especially in aquatic ecosystems, but prior studies have largely examined eDNA in cool temperate settings. By contrast, this study employs eDNA to survey the fish fauna in tropical Lake Bacalar (Mexico) with the additional goal of assessing the possible presence of invasive fishes, such as Amazon sailfin catfish and tilapia. Sediment and water samples were collected from eight stations in Lake Bacalar on three occasions over a 4-month interval. Each sample was stored in the presence or absence of lysis buffer to compare eDNA recovery. Short fragments (184–187 bp) of the cytochrome c oxidase I (COI) gene were amplified using fusion primers and then sequenced on Ion Torrent PGM or S5 before their source species were determined using a custom reference sequence database constructed on BOLD. In total, eDNA sequences were recovered from 75 species of vertebrates including 47 fishes, 15 birds, 7 mammals, 5 reptiles, and 1 amphibian. Although all species are known from this region, six fish species represent new records for the study area, while two require verification. Sequences for five species (2 birds, 2 mammals, 1 reptile) were only detected from sediments, while sequences from 52 species were only recovered from water. Because DNA from the Amazon sailfin catfish was not detected, we used a mock eDNA experiment to confirm our methods would enable its detection. In summary, we developed protocols that recovered eDNA from tropical oligotrophic aquatic ecosystems and confirmed their effectiveness in detecting fishes and diverse species of vertebrates.
Environmental DNA (eDNA) has gained popularity for biomonitoring, especially for the detection of invasive species and for baseline surveys of animal and plant communties [1,2]. This eDNA derives from cells shed into the environment as mucus, urine, feces, or gametes [2–7]. More than 120 articles have now considered eDNA, including a special issue on the topic . The rapid adoption of eDNA-based bioassessments reflects two factors: improved access to the reference sequences [9,10] required to link the short reads from eDNA to their source species, and the availability of high-throughput sequencers that can generate large volumes of data at modest cost.
The major goal of this study was to evaluate the applicability of eDNA for biomonitoring fishes in a tropical oligotrophic lake. Although eDNA has often been thought to degrade rapidly, factors such as temperature, alkalinity, and trophic state [31,62] affect its stability. For example, cold temperatures, low UV-B levels, and high pH slow eDNA degradation , while acidity promotes it [62,63]. The overall probability of eDNA detection also depends on its production which may vary by species, by season, by density, and diet, and its loss from the study system via water discharge or diffusion . The impacts of increased temperature on eDNA recovery is inconsistent. For example, a laboratory study showed that eDNA degradation increased with rising temperature, particularly in water samples from an oligotrophic lake . By contrast, Robson et al  evaluated effects of high water temperature and fish density on the detection of invasive Mozambique tilapia in ponds via eDNA protocols and found that increased water temperatures did not affect degradation rates. However, they did detect increased rates of eDNA shedding at 35°C. In our study we detected two non-native tilapia species (O. mossambicus and O. niloticus) in Lake Bacalar (S1 and S2 Tables). Sampling methodology and sample handling are crucial for eDNA preservation. Therefore, samples were preserved in the presence or absence of buffer to compare overall DNA yield and eDNA recovery.
We developed field sampling protocols and a HTS pipeline which enabled the efficient recovery of eDNA from several tropical aquatic ecosystems. Water samples consistently revealed more vertebrate species than sediment samples although about 10% of the species were only recovered from sediments.