2.3. Metals and Microbial Analyses
Metals Analyses . All sediment and soil samples were tested for
arsenic, cadmium, chromium, copper, lead, and nickel because of the
presence of nearby Superfund sites that exhibit the six metals and the
abundance of industries with discharge permits for these metals, located
upstream and within the HSC-GB system. For metals, digestion and
analysis followed EPA SW-846-6010D (U.S. EPA, 2014) 8,
and EPA 3050B (EPA, 1996), respectively (all of the analyses were
conducted by an EPA accredited lab). All bed sediment samples were
analyzed for TOC and grain size distribution within the 28-day limit for
hold time following the Lloyd Kahns (Kahn, 1988) and ASTM D4464 (ASTM,
2015) standard methods, respectively. For all bed sediment sampling
locations, field water quality parameters including pH, DO, and salinity
for the bed layer (0.3 meters above the bottom of the waterbody) were
measured using a field multi parameter water quality Sonde (600 XLM, YSI
Inc.). Table S1 in the Supplementary Information (SI) provides a list of
the sampled stations and their measured water quality parameters.
Trace metal concentrations in deposited soil samples were compared to
protective concentration levels (PCLs) from the Texas Risk Reduction
Program (TRRP) (TRRP, 2017) and regional screening levels (RSLs) for
residential and commercial soils from the U.S.EPA (U.S. EPA, 2017) (see
Table S2 in the SI). For bed sediments, combined PCLs (assuming metal
intake is through all possible pathways) from TRRP for sediments were
used (see Table S3 in the SI) in the comparison.
Microbial Analyses. Microbial genomic DNA was isolated from 250
mg of sediments using DNeasy Powersoil kit from Qiagen® (Germantown,
MD). Sequencing of the 16s rRNA v3, v4 and v5 region was completed in an
accredited lab (Genewiz®, New Jersey, USA) using the 16S MetaVx
Environmental protocol on Illumina MiSeq with 2x250 bp configuration
(the primer sequences used in the 16S MetaVx environmental sequencing
service are proprietary). Microbiome analyses were primarily carried out
using a QIIME2 (Caporaso et al. , 2010) pipeline. The minimum
nucleotide Phred quality score was set to 30; readings smaller or larger
than 400 nucleotides were eliminated using Deblur plugins in QIIME2.
Chimeric reads, including borderline sequences were removed via UCHIME
(Edgar et al. , 2011). A total of approximately 10,074,932 reads
were generated from the eleven samples. The Illumina sequence data have
been deposited in the NCBI Sequence Read Archive under accession number
PRJNA554335.