From Land to Lake: Contrasting Microbial Processes Across a Great Lakes Gradient of Organic Carbon and Inorganic Nutrient Inventories

Freshwater ecosystems have strong linkages to the terrestrial landscapes that surround them, and contributions of carbon and inorganic nutrients from soil, vegetation and anthropogenic sources subsidize autochthonous water body productivity to varying degrees. Abundant freshwater phytoplankton and bacterioplankton are key to linking the planet\u27s geosphere and atmosphere to the food webs in the hydrosphere through their growth and respiration. Rich resources that move through land margin waterways make them active sites for cycling organic carbon and thus important, but understudied, contributors to global climate. During 2010-2011, we examined seasonal changes in carbon and nutrient inventories, plankton community composition and metabolism along a land-to-lake gradient in a major West Michigan watershed at four interconnected habitats ranging from a small creek to offshore Lake Michigan. In all seasons Lake Michigan had significantly lower concentrations of CDOM and DOC than any of the other sites. Lake levels of NO3 were not significantly lower than tributaries other than Cedar Creek, and SRP was not measurable in any of the sites other than Cedar Creek. Bacterial production as % of GPP revealed a distinct land-to-lake gradient from an average of 448% in Cedar creek to 5% in Lake Michigan. Microbial activity in Cedar Creek (bacterial production 3- 93 μg C/L/d, and plankton respiration 9-193 μg C/L/d) was generally higher than all other sites. Muskegon Lake dominated GPP among the sites reaching a peak of \u3e1000 μg carbon/L/d during a large fall Microcystis bloom. Offshore Lake Michigan had less variation in GPP and R than the other sites with GPP:R ratio close to 1 in all seasons but spring. Metabolism appears to be substantially subsidized by terrigenous inputs in the creek/river ecosystem with heterotrophy dominant over autotrophy. Autotrophy was maximized in the coastal/estuary, whereas both autotrophy and heterotrophy were minimal but in near-balance in offshore waters receiving little subsidy from the land. Along this land-to-lake gradient terrestrial subsidies combined with a host of other factors making conditions “just right” for a hot-spot to emerge, highlighting Muskegon Lake estuary a “Goldilocks Zone” of net biological productivity

High levels of sewage contamination released from urban areas after storm events: A quantitative survey with sewage specific bacterial indicators

Background Past studies have demonstrated an association between waterborne disease and heavy precipitation, and climate change is predicted to increase the frequency of these types of intense storm events in some parts of the United States. In this study, we examined the linkage between rainfall and sewage contamination of urban waterways and quantified the amount of sewage released from a major urban area under different hydrologic conditions to identify conditions that increase human risk of exposure to sewage. Methods and findings Rain events and low-flow periods were intensively sampled to quantify loads of sewage based on two genetic markers for human-associated indicator bacteria (human Bacteroides and Lachnospiraceae). Samples were collected at a Lake Michigan estuary and at three river locations immediately upstream. Concentrations of indicators were analyzed using quantitative polymerase chain reaction (qPCR), and loads were calculated from streamflow data collected at each location. Human-associated indicators were found during periods of low flow, and loads increased one to two orders of magnitude during rain events from stormwater discharges contaminated with sewage. Combined sewer overflow (CSO) events increased concentrations and loads of human-associated indicators an order of magnitude greater than heavy rainfall events without CSO influence. Human-associated indicator yields (load per km2 of land per day) were related to the degree of urbanization in each watershed. Contamination in surface waters were at levels above the acceptable risk for recreational use. Further, evidence of sewage exfiltration from pipes threatens drinking water distribution systems and source water. While this study clearly demonstrates widespread sewage contamination released from urban areas, a limitation of this study is understanding human exposure and illness rates, which are dependent on multiple factors, and gaps in our knowledge of the ultimate health outcomes. Conclusions With the prediction of more intense rain events in certain regions due to climate change, sewer overflows and contamination from failing sewer infrastructure may increase, resulting in increases in waterborne pathogen burdens in waterways. These findings quantify hazards in exposure pathways from rain events and illustrate the additional stress that climate change may have on urban water systems. This information could be used to prioritize efforts to invest in failing sewer infrastructure and create appropriate goals to address the health concerns posed by sewage contamination from urban areas

Sewage reflects the microbiomes of human populations

© The Author(s), 2015. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in mBio 6 (2015): e02574-14, doi:10.1128/mBio.02574-14.Molecular characterizations of the gut microbiome from individual human stool samples have identified community patterns that correlate with age, disease, diet, and other human characteristics, but resources for marker gene studies that consider microbiome trends among human populations scale with the number of individuals sampled from each population. As an alternative strategy for sampling populations, we examined whether sewage accurately reflects the microbial community of a mixture of stool samples. We used oligotyping of high-throughput 16S rRNA gene sequence data to compare the bacterial distribution in a stool data set to a sewage influent data set from 71 U.S. cities. On average, only 15% of sewage sample sequence reads were attributed to human fecal origin, but sewage recaptured most (97%) human fecal oligotypes. The most common oligotypes in stool matched the most common and abundant in sewage. After informatically separating sequences of human fecal origin, sewage samples exhibited ~3× greater diversity than stool samples. Comparisons among municipal sewage communities revealed the ubiquitous and abundant occurrence of 27 human fecal oligotypes, representing an apparent core set of organisms in U.S. populations. The fecal community variability among U.S. populations was significantly lower than among individuals. It clustered into three primary community structures distinguished by oligotypes from either: Bacteroidaceae, Prevotellaceae, or Lachnospiraceae/Ruminococcaceae. These distribution patterns reflected human population variation and predicted whether samples represented lean or obese populations with 81 to 89% accuracy. Our findings demonstrate that sewage represents the fecal microbial community of human populations and captures population-level traits of the human microbiome.Funding for this work was provided by the NIH grant R01AI091829-01A1 to S.L.M. and M.L.S

Systematically Variable Planktonic Carbon Metabolism Along a Land-To-Lake Gradient in a Great Lakes Coastal Zone

During the summers of 2002–2013, we measured rates of carbon metabolism in surface waters of six sites across a land-to-lake gradient from the upstream end of drowned river-mouth Muskegon Lake (ML) (freshwater estuary) to 19 km offshore in Lake Michigan (LM) (a Great Lake). Despite considerable inter-year variability, the average rates of gross production (GP), respiration (R) and net production (NP) across ML (604 ± 58, 222 ± 22 and 381 ± 52 µg C L−1 day−1, respectively) decreased steeply in the furthest offshore LM site (22 ± 3, 55 ± 17 and −33 ± 15 µg C L−1day−1, respectively). Along this land-to-lake gradient, GP decreased by 96 ± 1%, whereas R only decreased by 75 ± 9%, variably influencing the carbon balance along this coastal zone. All ML sites were consistently net autotrophic (mean GP:R = 2.7), while the furthest offshore LM site was net heterotrophic (mean GP:R = 0.4). Our study suggests that pelagic waters of this Great Lakes coastal estuary are net carbon sinks that transition into net carbon sources offshore. Reactive and dynamic estuarine coastal zones everywhere may contribute similarly to regional and global carbon cycles

Chronicles of hypoxia: Time-series buoy observations reveal annually recurring seasonal basin-wide hypoxia in Muskegon Lake – A Great Lakes estuary

We chronicled the seasonally recurring hypolimnetic hypoxia in Muskegon Lake – a Great Lakes estuary over 3 years, and examined its causes and consequences. Muskegon Lake is a mesotrophic drowned river mouth that drains Michigan\u27s 2nd largest watershed into Lake Michigan. A buoy observatory tracked ecosystem changes in the Muskegon Lake Area of Concern (AOC), gathering vital time-series data on the lake\u27s water quality from early summer through late fall from 2011 to 2013 (www.gvsu.edu/buoy). Observatory-based measurements of dissolved oxygen (DO) tracked the gradual development, intensification and breakdown of hypoxia (mild hypoxia b4 mg DO/L, and severe hypoxia b2 mg DO/L) below the ~6 m thermocline in the lake, occurring in synchrony with changes in temperature and phytoplankton biomass in the water column during July–October. Time-series data suggest that proximal causes of the observed seasonal hypolimnetic DO dynamics are stratified summer water-column, reduced wind-driven mixing, longer summer residence time, episodic intrusions of cold DO-rich nearshore Lake Michigan water, nutrient run off from watershed, and phytoplankton blooms. Additional basin-wide water-column profiling (2011–2012) and ship-based seasonal surveys (2003–2013) confirmed that bottom water hypoxia is an annually recurring lake-wide condition. Volumetric hypolimnetic oxygen demand was high (0.07–0.15 mg DO/Liter/day) and comparable to other temperate eutrophic lakes. Over 3 years of intense monitoring, ~9–24% of Muskegon Lake\u27s volume experienced hypoxia for ~29–85 days/year – with the potential for hypolimnetic habitat degradation and sediment phosphorus release leading to further eutrophication. Thus, time-series observatories can provide penetrating insights into the inner workings of ecosystems and their external drivers

Urban microbial ecology of a freshwater estuary of Lake Michigan

Abstract Freshwater estuaries throughout the Great Lakes region receive stormwater runoff and riverine inputs from heavily urbanized population centers. While human and animal feces contained in this runoff are often the focus of source tracking investigations, non-fecal bacterial loads from soil, aerosols, urban infrastructure, and other sources are also transported to estuaries and lakes. We quantified and characterized this non-fecal urban microbial component using bacterial 16S rRNA gene sequences from sewage, stormwater, rivers, harbor/estuary, and the lake surrounding Milwaukee, WI, USA. Bacterial communities from each of these environments had a distinctive composition, but some community members were shared among environments. We used a statistical biomarker discovery tool to identify the components of the microbial community that were most strongly associated with stormwater and sewage to describe an “urban microbial signature,” and measured the presence and relative abundance of these organisms in the rivers, estuary, and lake. This urban signature increased in magnitude in the estuary and harbor with increasing rainfall levels, and was more apparent in lake samples with closest proximity to the Milwaukee estuary. The dominant bacterial taxa in the urban signature were Acinetobacter, Aeromonas, and Pseudomonas, which are organisms associated with pipe infrastructure and soil and not typically found in pelagic freshwater environments. These taxa were highly abundant in stormwater and sewage, but sewage also contained a high abundance of Arcobacter and Trichococcus that appeared in lower abundance in stormwater outfalls and in trace amounts in aquatic environments. Urban signature organisms comprised 1.7% of estuary and harbor communities under baseflow conditions, 3.5% after rain, and >10% after a combined sewer overflow. With predicted increases in urbanization across the Great Lakes, further alteration of freshwater communities is likely to occur with potential long term impacts on the function of estuarine and nearshore ecosystems

High levels of sewage contamination released from urban areas after storm events: A quantitative survey with sewage specific bacterial indicators.

BACKGROUND:Past studies have demonstrated an association between waterborne disease and heavy precipitation, and climate change is predicted to increase the frequency of these types of intense storm events in some parts of the United States. In this study, we examined the linkage between rainfall and sewage contamination of urban waterways and quantified the amount of sewage released from a major urban area under different hydrologic conditions to identify conditions that increase human risk of exposure to sewage. METHODS AND FINDINGS:Rain events and low-flow periods were intensively sampled to quantify loads of sewage based on two genetic markers for human-associated indicator bacteria (human Bacteroides and Lachnospiraceae). Samples were collected at a Lake Michigan estuary and at three river locations immediately upstream. Concentrations of indicators were analyzed using quantitative polymerase chain reaction (qPCR), and loads were calculated from streamflow data collected at each location. Human-associated indicators were found during periods of low flow, and loads increased one to two orders of magnitude during rain events from stormwater discharges contaminated with sewage. Combined sewer overflow (CSO) events increased concentrations and loads of human-associated indicators an order of magnitude greater than heavy rainfall events without CSO influence. Human-associated indicator yields (load per km2 of land per day) were related to the degree of urbanization in each watershed. Contamination in surface waters were at levels above the acceptable risk for recreational use. Further, evidence of sewage exfiltration from pipes threatens drinking water distribution systems and source water. While this study clearly demonstrates widespread sewage contamination released from urban areas, a limitation of this study is understanding human exposure and illness rates, which are dependent on multiple factors, and gaps in our knowledge of the ultimate health outcomes. CONCLUSIONS:With the prediction of more intense rain events in certain regions due to climate change, sewer overflows and contamination from failing sewer infrastructure may increase, resulting in increases in waterborne pathogen burdens in waterways. These findings quantify hazards in exposure pathways from rain events and illustrate the additional stress that climate change may have on urban water systems. This information could be used to prioritize efforts to invest in failing sewer infrastructure and create appropriate goals to address the health concerns posed by sewage contamination from urban areas

EcoBLMcrX, a classical modification-dependent restriction enzyme in <i>Escherichia coli</i> B: Characterization <i>in vivo</i> and <i>in vitro</i> with a new approach to cleavage site determination

<div><p>Here we characterize the <u>m</u>odification-<u>d</u>ependent restriction <u>e</u>nzyme (MDE) EcoBLMcrX <i>in vivo</i>, <i>in vitro</i> and in its genomic environment. MDE cleavage of modified DNAs protects prokaryote populations from lethal infection by bacteriophage with highly modified DNA, and also stabilizes lineages by reducing gene import when sparse modification occurs in the wrong context. The function and distribution of MDE families are thus important. Here we describe the properties of EcoBLMcrX, an enzyme of the <i>E</i>. <i>coli B</i> lineage, <i>in vivo</i> and <i>in vitro</i>. Restriction <i>in vivo</i> and the genome location of its gene, <i>ecoBLmcrX</i>, were determined during construction and sequencing of a B/K-12 hybrid, ER2566. In classical restriction literature, this B system was named <i>r</i>_<i>6</i> or <i>rglA</i>_<i>B</i>. Like many genome defense functions, <i>ecoBLmcrX</i> is found within a genomic island, where gene content is variable among natural <i>E</i>. <i>coli</i> isolates. <i>In vitro</i>, EcoBLMcrX was compared with two related enzymes, BceYI and NhoI. All three degrade fully cytosine-modified phage DNA, as expected for EcoBLMcrX from classical T4 genetic data. A new method of characterizing MDE specificity was developed to better understand action on fully-modified targets such as the phage that provide major evolutionary pressure for MDE maintenance. These enzymes also cleave plasmids with ^m5C in particular motifs, consistent with a role in lineage-stabilization. The recognition sites were characterized using a site-ranking approach that allows visualization of preferred cleavage sites when fully-modified substrates are digested. A technical constraint on the method is that ligation of one-nucleotide 5' extensions favors G:C over A:T approximately five-fold. Taking this bias into account, we conclude that EcoBLMcrX can cleave 3' to the modified base in the motif R^m5C|. This is compatible with, but less specific than, the site reported by others. Highly-modified site contexts, such as those found in base-substituted virulent phages, are strongly preferred.</p></div

High levels of sewage contamination released from urban areas after storm events: A quantitative survey with sewage specific bacterial indicators - Fig 2

<p>(A) Streamflow (upper panel) and corresponding HB, Lachno2, ruminant, <i>E</i>. <i>coli</i>, and enterococci indicator concentrations (lower panel) measured during a combined sewer overflow in the Milwaukee estuary in Milwaukee, Wisconsin from 4/9/2015–4/13/2015 and (B) streamflow (upper panel) and corresponding HB, Lachno2, ruminant, <i>E</i>. <i>coli</i>, and enterococci indicator concentrations (lower panel) measured during two rain events in the Milwaukee estuary in Milwaukee, Wisconsin from 6/11/2015–6/13/2015 (event 8) and 6/14 /2015–6/16/15 (event 9). Vertical black dashed lines represent the beginning and ending dates and times that were defined for each event. HB, human <i>Bacteroides</i>; Lachno2, human Lachnospiraceae.</p

Control WebLogo—Type II enzyme on unmodified target.

<p>Control WebLogo—Type II enzyme on unmodified target.</p