Pharmaceutical effects on biofilm functioning quantified via contaminant exposure substrates presented in Rosi et al. 2018.

An ongoing component of the Baltimore urban long-term ecological research (LTER) project (Baltimore Ecosystem Study, BES) is the use of the watershed approach and monitoring of stream water quality to evaluate the impacts of multiple chemical stressors on urban stream ecosystem functioning within Baltimore. The LTER research has focused on the Gwynns Falls watershed, which spans a gradient from highly urban, urban-residential, and suburban zones. In addition, a forested watershed serves as a reference. The long-term sampling network includes four longitudinal sampling sites along the Gwynns Falls mainstem, as well as several small (40-100 ha) watershed within or near the Gwynns Falls, providing data on water quality in different land use zones of the watersheds. Each study site is continuously monitored for discharge and is sampled weekly for water chemistry. Those data are available elsewhere on the BES website. We are interested in studying the effects of pharmaceuticals and personal care products (PPCPs) on stream biofilm functioning within urban streams. We constructed and deployed contaminant exposure substrates (CES) in four streams within the greater Baltimore, Maryland, USA area to measure the responses of biofilms in each of the four streams to caffeine, ciprofloxacin, cimetidine, diphenhydramine, and no PPCPs. We incubated CES for 2 weeks in the four different streams, with five replicate CES for each treatment being randomly placed on plastic L-bars within each stream. After the two-week deployment, we used the light-dark incubation approach to estimate gross primary production and community respiration via change in dissolved oxygen (DO) for biofilms colonizing substrates (either cellulose sponges to select heterotrophic biofilms or fritted glass disks to select for autotrophic biofilms) topping each CES. This dataset includes all of the raw data for the light-dark incubations used to calculate gross primary production and community respiration for the various CES treatments at different sites. A subset of these data were used to estimate the effects of PPCPs on community respiration that is presented in the following publication: Rosi, E.J., H.A. Bechtold, D. Snow, M. Rojas, A.J. Reisinger, and J.J. Kelly. 2018. Urban stream microbial communities show resistance to pharmaceutical exposure. Ecosphere 9(1):e02041. doi: 10.1002/ecs2.2041 Each stream studied in this dataset is a core BES monitoring stream, and additional water chemistry and hydrology data are available elsewhere on the BES data portal. Codes and abbreviations 1 - GwynnsRun - Gwynns Run at Gwynns Falls - Urban 2 - GwynnsFalls - Gwynns Falls at Carroll Park - Urban 3 - DeadRun - Dead Run at Krome Avenue - Urban 4 - GwynnsBrook - Gwynns Falls at Gwynnbrook Avenue (Delight) - Suburban Column,Column Name,Variable-if different than Column Name (units) A,cup,identifier for CES cup replicate B,site, Site C,compound, Treatment - can be one of: Ciprofloxacin, Cimetidine, Caffeine, Control, or Diphenhydramine D,substrate, substrate topping the CES used to select for either autotrophic or heterotrophic biofilms - can be either Cellulose (heterotrophic) or Fr. Glass (autotrophic) E,light, light treatment for the incubation, either light (used to estimate NPP and GPP) or dark (used to estimate CR) F,o2.init, initial dissolved oxygen (DO) concentration of incubation water (mg O2/L) G,o2.final, final DO concentration after CES incubation has completed (mg O2/L) H,t.start, start time for the incubation - time when incubation vials were filled and closed (HH:MM:SS) I,t.end, end time for the incubation - time when incubation vials were opened and o2.final was measured (HH:MM:SS) J,blank.o2.init, initial DO concentration of incubation water used for blank incubation vials (no CES substrate) that are used to correct for water column activity (mg O2/L) K,blank.o2.final, final DO concentration of water in blank incubations that are used to correct for water column activity (mg O2/L) L,blank.t.start, start time for the blank incubations (HH:MM:SS) M,blank.t.end, end time for the blank incubations (HH:MM:SS) Methods: We used CES to measure the responses of biofilms in each of the four streams to caffeine, cimetidine, ciprofloxacin, and diphenhydramine. This contaminant exposure method allowed us to test the effects of PPCPs on the structure and function of microbial communities in situ. CES consisted of 30-mL polyethylene cups filled with a 2% (by weight) agarose gel amended with caffeine, ciprofloxacin, cimetidine, diphenhydramine, or no pharmaceutical. The concentration of PPCP in each cup was 0.05 mol/L. Each cup was capped with either a cellulose sponge cloth or a fritted glass disk to promote colonization by heterotrophic and autotrophic biofilms, respectively. Five replicate CES of each PPCP treatment and an additional control treatment were secured to the stream bottom on plastic L-bars, with treatments and replicates arranged randomly. CES were deployed for two weeks in March 2012. After deployment, CES were collected and transported to the laboratory on ice. We used the light-dark incubation approach to estimate GPP and CR from the different substrates for each CES. Each substrate was placed ina 50-mL centrifuge tube and each tube was filled with filtered stream water with known initial DO concentration. Each tube was capped underwater to remove all air bubbles and was incubated in the light and dark for 2-4h for each light treatment. We included blank tubes, which were filled with filtered stream water only, to correct for changes in background DO. After the 2-4h incubation period, we measured the final DO concentration in each of the tubes. These incubation data are what are included in this dataset. To calculate GPP and CR, standard light-dark DO incubation methods can be followed. For example, see: Tank, J.L., A.J. Reisinger, and E.J. Rosi. 2017. Ch. 31: Nutrient limitation and uptake. In Methods in Stream Ecology Vol 2: Ecosystem Function (3rd Edition), eds: G.A. Lamberti and F.R. Hauer. Academic Press, San Diego, CA, USA.

巴尔的摩城市长期生态研究（Long-Term Ecological Research, LTER）项目（又称巴尔的摩生态系统研究（Baltimore Ecosystem Study, BES））的持续组成部分，采用流域方法与河流水质监测，评估多种化学胁迫因子对巴尔的摩市内城市溪流生态系统功能的影响。该LTER研究聚焦于格温斯福尔斯流域，该流域涵盖高度城市化、城市居住及郊区的土地利用梯度；此外还设置了一处森林流域作为对照参考。长期采样网络包含沿格温斯福尔斯干流的4个纵向采样点位，以及格温斯福尔斯流域内或邻近的数个小型（40-100公顷）子流域，可为不同土地利用分区的流域水质提供监测数据。每个研究点位均持续监测径流量，并每周采集水化学样品，相关数据可通过BES官网获取。

本研究关注药物与个人护理产品（Pharmaceuticals and Personal Care Products, PPCPs）对城市溪流生物膜功能的影响。我们在美国马里兰州大巴尔的摩地区的4条溪流中构建并部署了污染物暴露底物（Contaminant Exposure Substrates, CES），以测定4条溪流中生物膜分别对咖啡因、环丙沙星、西咪替丁、苯海拉明及无PPCPs对照的响应。

我们将CES在4条溪流中孵育2周，每个处理设置5个重复CES，随机放置于各溪流内的塑料L型支架上。为期2周的部署结束后，我们采用明暗孵育法，通过测定附着于底物（用于培养异养生物膜的纤维素海绵，或用于培养自养生物膜的多孔玻璃圆盘）上的生物膜的溶解氧（Dissolved Oxygen, DO）变化，估算总初级生产力（Gross Primary Production, GPP）与群落呼吸（Community Respiration, CR）。

本数据集包含所有用于计算各点位不同CES处理下GPP与CR的明暗孵育原始数据。其中一部分数据被用于评估PPCPs对群落呼吸的影响，相关结果发表于以下论文：
Rosi, E.J., H.A. Bechtold, D. Snow, M. Rojas, A.J. Reisinger, and J.J. Kelly. 2018. Urban stream microbial communities show resistance to pharmaceutical exposure. Ecosphere 9(1):e02041. doi: 10.1002/ecs2.2041
本数据集涉及的每条研究溪流均为BES核心监测溪流，额外的水化学与水文数据可通过BES数据门户获取。

代码与缩写说明
1 - GwynnsRun：格温斯福尔斯的格温斯溪（Gwynns Run at Gwynns Falls），城市型
2 - GwynnsFalls：卡罗尔公园的格温斯福尔斯（Gwynns Falls at Carroll Park），城市型
3 - DeadRun：克罗姆大道的死溪（Dead Run at Krome Avenue），城市型
4 - GwynnsBrook：格温斯布鲁克大道的格温斯福尔斯（德尔莱特）（Gwynns Falls at Gwynnbrook Avenue (Delight)），郊区型

列号、列名、变量说明（若与列名不同，含单位）
A, cup：CES杯重复样本标识符
B, site：采样位点
C, compound：处理因子，可选值为：环丙沙星（Ciprofloxacin）、西咪替丁（Cimetidine）、咖啡因（Caffeine）、对照（Control）或苯海拉明（Diphenhydramine）
D, substrate：CES顶部的培养底物，用于筛选异养或自养生物膜，可选值为：纤维素（Cellulose，异养型）或多孔玻璃（Fr. Glass，自养型）
E, light：孵育光照处理，可选“光照”（用于估算净初级生产力NPP与总初级生产力GPP）或“黑暗”（用于估算群落呼吸CR）
F, o2.init：孵育用水的初始溶解氧（DO）浓度（mg O2/L）
G, o2.final：CES孵育结束后的最终溶解氧（DO）浓度（mg O2/L）
H, t.start：孵育开始时间——孵育管密封装填时的时间（HH:MM:SS）
I, t.end：孵育结束时间——孵育管开启并测定o2.final时的时间（HH:MM:SS）
J, blank.o2.init：空白孵育管（无CES底物）的初始溶解氧（DO）浓度，用于校正水柱本底活性（mg O2/L）
K, blank.o2.final：空白孵育管的最终溶解氧（DO）浓度，用于校正水柱本底活性（mg O2/L）
L, blank.t.start：空白孵育的开始时间（HH:MM:SS）
M, blank.t.end：空白孵育的结束时间（HH:MM:SS）

研究方法
我们采用CES测定4条溪流中生物膜分别对咖啡因、西咪替丁、环丙沙星及苯海拉明的响应。该污染物暴露方法可原位测试PPCPs对微生物群落结构与功能的影响。CES由30mL聚乙烯杯制成，内部填充质量分数为2%的琼脂糖凝胶，并添加咖啡因、环丙沙星、西咪替丁、苯海拉明或不添加任何药物，其中每杯内PPCP的浓度为0.05mol/L。每个杯子分别以纤维素海绵布或多孔玻璃圆盘作为顶盖，分别促进异养与自养生物膜的定植。每个PPCP处理及额外的对照处理均设置5个重复CES，随机固定于溪流底部的塑料L型支架上。CES于2012年3月部署，为期2周。部署结束后，收集CES并置于冰上运输至实验室。

我们采用明暗孵育法，针对每个CES的不同底物估算GPP与CR。将每个底物放置于50mL离心管中，加入已知初始DO浓度的过滤溪流河水，水下封管以去除所有气泡，分别进行光照与黑暗孵育2-4小时。设置仅加入过滤溪流河水的空白管，用于校正背景DO变化。孵育2-4小时后，测定每个管内的最终DO浓度。本数据集包含的即为该孵育实验的相关数据。总初级生产力与群落呼吸的计算可遵循标准的明暗DO孵育方法，例如参考Tank等人2017年的章节：
Tank, J.L., A.J. Reisinger, and E.J. Rosi. 2017. Ch. 31: Nutrient limitation and uptake. In Methods in Stream Ecology Vol 2: Ecosystem Function (3rd Edition), eds: G.A. Lamberti and F.R. Hauer. Academic Press, San Diego, CA, USA.