What’s all that racket! Soundscapes, phenology, and biodiversity in estuaries

change rapidly or gradually in response to environmental changes and human use; thus, it has potential to provide a measure of resilience or shifting baselines in a globally changing environment. Materials and methods Study area The May River (32˚12’49”N, 80˚52’23”W) is located inland of the southern SC coast (Fig 5). This large subtidal river is ~22 km long and ~0.01 km wide near the source, and ~1 km wide at the mouth. The river is bordered by extensive patches of smooth cordgrass and oyster reefs (i.e. eastern oyster) with the town of Bluffton on the north-eastern side and Hilton Head Island at the mouth of the river. Water depth ranges from ~3 to ~7 m near the source and from ~4 to ~18 m near the mouth depending upon large semidiurnal tides. This area experiences a humid subtropical climate with hot summers and mild winters. Data collection and analysis We deployed DSG-Ocean recorders (Loggerhead Instruments, Sarasota, FL, USA), water level and temperature loggers (HOBO 100-Foot Depth Water Level Data Logger U20-001-02-Ti and HOBO Water Temperature Pro v2 U22-001, Onset Computer Corporation, Bourne, MA, USA) at stations 9M, 14M, and 37M between February 2013 and December 2018 following methods previously described (Fig 5) [28]. Recorders collected sound samples for 2 min every 20 min at a sampling rate of 80 kHz over 22 deployments that were approximately 90 days long. We determined the root mean square (rms) SPL for the entire data set (i.e. each 2 min wav file every 20 min; 392,106 files for all 3 stations) for high (i.e. 7000–40,000 Hz), low (i.e. 50–

1200 Hz), and broadband (i.e. 1–40,000 Hz) frequencies using custom scripts created in MATLAB R2017b (MathWorks, Inc., Natick, MA, USA). We chose these ranges based on pre- vious studies that revealed specific call frequencies for black drum (70–90 Hz), silver perch (1000–1280 Hz), oyster toadfish (190–200 Hz), spotted seatrout (200–270 Hz), red drum (120–160 Hz), and snapping shrimp (50–40 kHz) [28]. In the May River estuary, previous studies have discovered that the soundscape is composed mainly of biological sounds (i.e. snapping shrimp, fish, and bottlenose dolphins), physical sounds (i.e. wave, wind, water flow, and rain), and anthropogenic noise (i.e. recreational boats) [18]. Broadband frequency SPL values reflected biological sounds (i.e. snapping shrimp, fish, and bottlenose dolphins), physi- cal sounds (i.e. wave, wind, water flow, and rain), and anthropogenic noise (i.e. recreational boats) [18]. Low frequency SPLs included fish calls, the lower bandwidth of snapping shrimp snaps, bottlenose dolphin vocalizations (which were few and random), physical sounds, and anthropogenic noise [18]. High frequency SPLs included snapping shrimp snaps, high fre- quency vocalizations of bottlenose dolphins, physical sounds, and anthropogenic noise [18]. In order to decipher biological patterns from physicals sounds and noise, we subsampled the data set and manually analyzed files recorded on the hour (i.e. 130,702 files for all 3 sta- tions). We flagged the files that contained biological sounds (i.e. snapping shrimp, fish, and

bottlenose dolphin), physical sounds (i.e. wave, wind, water flow, and rain), and anthropogenic noise (i.e. recreational boats) [18]. Then, we removed the files that contained physical sounds and anthropogenic noise from the subsampled data set and reanalyzed high, low, and broad- band SPLs. This approach ensured that the patterns observed where of biological nature. We created heat maps in MATLAB R2017b using the entire data set representing all SPL values (i.e. 2 min on the hour including sounds of biological, physical, and anthropogenic origin) and the subsampled data set (i.e. 2 min on the hour including only sounds of biological origin). PLOS ONE Estuarine soundscapes and biodiversity PLOS ONE | https://doi.org/10.1371/journal.pone.0236874 September 3, 2020 9 / 18