Biodiversity is critical to ecosystem functioning and is threatened by anthropogenic activity

They allowed for a 1% chance of mutation of each experiment and component to allow for global search. They also discovered that the response space was multi-modal and had interactions between components, which confirmed the need for global optimization of fermentation and bio-processing problems.Microbes perform key ecosystem functions , necessitating a better understanding of how microbes and microbial diversity respond to environmental stressors . Streams are examples of threatened ecosystems, where watershed modification decreases stream integrity and water quality , altering macro-invertebrate, fish, and microbial diversity . The use of macro-invertebrate and fish indices to assess stream conditions is fundamental to stream ecology and depends on known relationships between stream integrity and community structure . The Benthic Index of Biotic Integrity is one such index, using the abundance and diversity of stream benthic macro-invertebrates to accurately distinguish degraded streams classified based on stream chemical and physical criteria . Biotic indices are calibrated to specific regions, as the distribution of stream macro-invertebrates is controlled by a combination of dispersal limitation and local environmental conditions . Despite their abundance and ecological importance, natural microbial communities, unlike macro-invertebrates, are not used in stream monitoring programs to assess stream conditions. As with macro-invertebrates, dispersion and environmental selection control the spatial distribution of microbes along stream continuums . Dispersion,growing raspberries in pots or the advection of microbes from the surrounding landscape, impacts head water stream community composition, and with increasing stream order, environmental sorting becomes more important as stream residence times increase .

Several studies have demonstrated the influence of the surrounding landscape on stream microbes, showing that watershed urbanization leads to shifts in bacterial communities . While alpha diversity generally remains constant , the abundances of taxa associated with anthropogenic activity and high-nutrient conditions increase in urbanized streams . Similar to larger organisms, microbes respond to environmental disturbance and are strongly influenced by watershed land use ; therefore, their distribution may be used to further characterize stream conditions. Microbes mediate important stream ecosystem functions, controlling the movement of carbon and nitrogen through freshwater ecosystems . Previous studies demonstrate the effects of urbanization on stream nutrient transformations, such as nitrogen uptake , nitrogen retention , and carbon processing . Community respiration determines the fate of terrestrial carbon in head water streams, where carbon is either lost as carbon dioxide during respiration or transported farther downstream . Community respiration is often used to assess ecosystem function , as rates are influenced by watershed land use , correlated with stream chemistry , and sensitive to pollutants . The effects of urbanization on stream dissolved organic matter quality and respiration have previously been demonstrated, and stream microbial community structure can potentially be used to monitor these ecosystem functions. In addition to respiration, dissolved organic matter fuels stream denitrification and the microbial reduction of nitrate to nitrous oxide and dinitrogen gases . Denitrification removes nitrogen from streams and is credited as the major source of the greenhouse gas N2O . Watershed land use and anthropogenic nitrogen loading alter rates of stream denitrification , increasing the amount of nitrogen transported downstream and emissions of N2O to the atmosphere . Urbanization has been linked to changes in denitrifier community composition , and a previous study linked changes in denitrifier composition to changes in denitrification potential, and therefore nitrogen loss, in urban streams .

However, it is less clear how changes in microbial community composition in response to land use modification alter N2O production. The goal of this study was to identify stream microbes that respond to watershed urbanization and agricultural development. These anthropogenic factors alter microbial diversity and community structure, which can be used to assess stream conditions and ecosystem functioning. We measured microbial diversity using 16S rRNA gene amplicon sequencing across 82 head water streams within the Chesapeake Bay watershed in the state of Maryland in the spring and summer for 2 years. Measurements were collected in conjunction with stream physicochemical parameters and a macro-invertebrate indicator of stream health. Additionally, at a subset of streams, water column and sediment community respiration were measured using oxygen consumption methods, and N2O concentrations were measured using gas chromatography. We determined how stream bacteria and archaea are distributed across gradients of watershed land use and stream conditions, assessed how changes in microbial community composition relate to benthic macro-invertebrate diversity and traditional indices of stream conditions, and determined how these changes influence stream function by relating microbial community composition to rates of microbial respiration and concentrations of N2O.The aims of this study were to understand how stream bacteria and archaea are distributed across gradients of watershed land use and water quality, to assess how changes in microbial community composition relate to benthic macro-invertebrate diversity, and to discern how changes in stream conditions alter stream ecosystem processes, as reflected in community respiration and N2O concentrations. Bacterial and archaeal diversity significantly differed across the geographic regions of Maryland , demonstrating the influence of the surrounding landscape on headwater stream microbial communities.

Regional alluvium composition likely influenced stream alpha diversity, causing lower alpha diversity in Coastal Plain streams . Sediments on the Coastal Plain of the eastern United States are composed of gravel, sand, silt, and clay , making streams more embedded . Embeddedness was the environmental factor that most strongly negatively correlated with Shannon diversity , and homogeneous fine sediments have been shown to have lower diversity than that of sites with riffles, shallow turbulent sections . Similarly, community structure varied across the geographic regions and strongly correlated with DOC concentration, pH, and embeddedness , all of which significantly differentiate Coastal Plain streams from the other regions . This finding is in agreement with those from previous studies, demonstrating the strong influence of DOC concentration and pH on freshwater communities . Despite the strong influence of stream chemistry on microbial communities , in this study, geodesic distance explained more of the variation in community composition than environmental distance. Partial Mantel test results indicated that community structure was correlated with geographic distance rather than the measured environmental variables. Geographic distance is likely a strong controlling factor in structuring head water stream communities because there are regional differences in landscape, and stream microbes are locally seeded from the surrounding soil . Alpha diversity was greatest in spring, when water flow through the landscape is greatest and, therefore, when advection of microbes from the surrounding landscape is greatest to head water streams. While seasonal changes in microbial diversity during fall and winter are unknown, the higher diversity in spring than in summer was likely due to higher terrestrial inputs in spring, further demonstrating the influence of landscape on stream microbial communities. Distance-decay relationships were also observed between water column and sediment community similarity and geodesic distance , further highlighting the finding that head water stream microbes display geographic distribution patterns. Alternatively,plant pot with drainage the distance-decay relationship could be a result of spatial differences in unmeasured environmental variables. Microbial distance-decay relationships have been observed previously in streams . Z-values represent the rate at which species similarity decreases with increasing distance; in this study, Z-values are similar to microbial values from soil, salt marshes, and lakes but lower than regional differences observed in salt marshes , suggesting different dispersal limitations across regional scales. In contrast to highly urban and agricultural streams, community dissimilarity in highly forested streams did not increase with distance. Neither geographic distance nor environmental distance correlated with community structure, implying that highly forested streams have a similar terrestrial microbial source. Microbial diversity differed in streams in watersheds with high urban, agricultural, and forested land use. In contrast to previous studies, degraded streams had lower alpha diversity than that of forested streams , likely due to elevated pollution and habitat loss. Several abundant and pervasive taxa found in urban and agricultural streams are often associated with high-nutrient and low-oxygen environments. Members of the order Burkholderialeswere abundant in urban streams and correlated strongly with several anthropogenic nutrients .

Comamonadaceae are often associated with high nutrient conditions and are ubiquitous in many environments, including aquatic, soil, activated sludge, and wastewater . Comamonadaceae have previously been associated with urban streams and have been found to have the highest number of urban-tolerant taxa . Sulfurospirillum spp., in the order Campylobacterales, were abundant in highly agricultural streams and are often associated with microaerophilic polluted habits, commonly growing on arsenate or selenate using NO3 and sulfur compounds as electron acceptors . In contrast, an unclassified and potentially phototrophic member of Acidobacteria and Hyphomicrobiaceaewere more abundant in forested streams. These taxa are often associated with low nutrient conditions and were previously identified as indicators of forested streams and shown to decrease in abundance with increasing watershed urbanization . Only weak associations were detected between sediment and water microbial community composition and B-IBI scores. This is in contrast to findings of Simoninet al., who found that stream microbial community structure correlates with a macro-invertebrate biotic index in North Carolina . Simonin et al. identified concurrent changes in microbial taxa and environmental conditions associated with the biotic index, finding a higher number of negatively responding taxa than positive responding taxa . Here, only one taxon was more abundant and pervasive in streams in good condition, while several taxa were found to be abundant and pervasive in streams in very poor condition . Hydrogenophaga spp. and an unclassified member of Desulfobacterales, commonly associated with anaerobic, reducing, and contaminated environments , were both more abundant in streams in very poor condition . Forested water communities were more even than agricultural and urban communities, suggesting that certain taxa increase in abundance disproportionality in degraded streams, which is likely why the indicator analysis identified more taxa in streams in very poor condition. The findings here suggest that land use cover and stream chemistry are better predictors of head water stream microbial community composition than are macro-invertebrate indices of stream conditions. In agreement with the idea that structure determines function, in this study, water community respiration correlated with microbial community composition. Similarly, previous studies report that changes in community metabolism, specifically, the degradation of organic matter, are related to shifts in community composition and diversity . In contrast, other studies report that respiration depends on substrate availability rather than community composition due to functional redundancy , finding no connection between stream bacterial diversity and the activity of enzymes associated with carbon cycling . The weaker correlation between community respiration and community composition in sediments compared to water samples could be due to a high level of functional redundancy within sediment communities; if dominated by generalists, shifts in community composition would likely not significantly affect rates of respiration . Degraded streams had lower rates of community respiration than forested streams, as evidenced by the positive correlation between water respiration and forest cover and the negative correlation between respiration and urban cover . Rates of community respiration also negatively correlated with several physicochemical variables , including conductivity, Cl , Ca, Mg, pH, SO4 2 , and ANC. All signatures of anthropogenic influence, ANC, Cl and Zn concentrations, and pH, were found to previously correlate with benthic stream respiration across the Highlands, Piedmont, and Coastal Plain regions of the eastern United States , and Zn is a common urban pollutant . These results suggest that environmental conditions associated with land use drive differences in community respiration, altering carbon processing in head water streams. In addition to altering carbon transformations, watershed modification affected stream nitrogen processing. Agricultural streams had higher N2O concentrations than those of forested streams , with higher N2O concentrations being associated with elevated TN, NO2 , NO3 , and NH4  concentrations . The N2O concentrations measured in this study, at 0.22 to 4.41 g N2O liter 1 , were comparable to the values reported for agricultural streams in Illinois but lower than values published for agricultural drainage waters in Scotland, UK and higher than values reported for other forested and agricultural streams . N2O production is known to vary by land use, with higher production from denitrification in streams in agricultural and urban basins , and changes in community composition have been shown to influence denitrification rates in agricultural and urban streams . However, Audet et al. found no difference in N2O concentrations measured in forested and agricultural streams in Sweden .