Benthic Fluxes of Fluorescent Dissolved Organic Material, Salt, and Heat Measured by Multiple-Sensor Aquatic Eddy Covariance.

Aquatic eddy covariance (AEC) is an in situ technique for measuring fluxes in marine and freshwater systems that is based on the covariance of velocity and concentration measurements. To date, AEC has mainly been applied to the measurement of benthic oxygen fluxes. Here, development of a fast multiple-channel sensor enables the use of AEC for measurement of benthic fluxes of fluorescent material, salt, and heat at three distinct sites in Massachusetts, USA, including the Connecticut River, the Concord River, and Upper Mystic Lake. Benthic fluxes of salt, useful as a tracer for groundwater input (submarine groundwater discharge), were consistent with independent measurements made with seepage meters. Eddy fluxes of heat were consistent with the balance of incoming solar radiation and thermal conduction at the sediment surface. Benthic eddy fluxes of fluorescent dissolved organic material (FDOM) revealed a substantial net downward flux in the humic-rich Concord River, suggesting that microbial consumption of dissolved organic carbon in the sediment was significant. Simultaneous measurement of several fluxes expands the utility of AEC as a biogeochemical tool while enabling checks for mutual consistency among data channels.

A case study in participatory science with mutual capacity building between university and tribal researchers to investigate drinking water quality in rural Maine.

BACKGROUND: Participatory science or citizen science is increasingly being recognized for providing benefits to scientists and community members alike. However, most participatory science projects include community researchers only in the sample collection phase of the research project. Here we describe how a rural tribal community and urban university utilized participatory science methods to engage community researchers across an entire research study, creating numerous opportunities for mutual capacity building. OBJECTIVES: Researchers from MIT and the Sipayik Environmental Department, a tribal government department, partnered to co-launch a participatory science project to analyze municipal and private well drinking water quality in households in three Maine communities. The objective was to provide households with information about metals, primarily lead and arsenic, in their drinking water, and to improve public education, community partnerships, and local scientific capacity. METHODS: MIT and Sipayik researchers engaged local communities through public community meetings, mailed flyers sent to residents, and meetings with local stakeholders. MIT and community researchers worked together to design and implement the study to quantify metals in community drinking water samples, as well as hold capacity-building trainings. Individual drinking water results were communicated to households, and generalized results were discussed at community meetings in the report-back phase. RESULTS: The study attained a 29% household participation rate in the region. The researchers completed the analysis and report-back on 652 water samples. Isolated incidences of lead and geologically-attributable arsenic exceeding EPA standards were found. Individual report-backs of the results enabled local participatory scientists to make their own informed public health decisions. The study produced methodologies for navigating potential ethical issues, working with diverse communities, and collaborating over challenging geographical distances. DISCUSSION: This project developed methodologies to build long-term relationships with local scientists and to engage community members and enhance the environmental literacy of rural communities. Both MIT and Sipayik researchers learned from each other throughout the project; Sipayik researchers built technical capacity while MIT researchers gained local and cultural understanding. Community outreach methods were most effective when sent directly to residents as mailed flyers or through Sipayik researchers' outreach.

Vertical transport of sediment-associated metals and cyanobacteria by ebullition in a stratified lake.

Bubbles adsorb and transport particulate matter both in industrial and marine systems. While methane-containing bubbles emitted from anoxic sediments are found extensively in aquatic ecosystems, relatively little attention has been paid to the possibility that such bubbles transport particle-associated chemical or biological material from sediments to surface waters of freshwater lakes. We quantified transport of particulate material from sediments to the surface by bubbles in Upper Mystic Lake, MA and in a 15 m tall experimental column. Vertical particle transport was positively correlated with the volume of gas bubbles released from the sediment. Particles transported by bubbles originated almost entirely in the sediment, rather than being scavenged from the water column. Concentrations of arsenic, chromium, lead, and cyanobacterial cells in bubble-transported particulate material were similar to those of bulk sediment, and particles were transported from depths exceeding 15 m, resulting in daily fluxes as large as 0.18 mg of arsenic m-2 and 2 × 104 cyanobacterial cells m-2 in the strongly stratified Upper Mystic Lake. While bubble-facilitated arsenic transport currently appears to be a modest component of total arsenic cycling in this lake, bubble-facilitated cyanobacterial transport could comprise as much as 17% of recruitment in this lake and may thus be of particular importance in large, deep, stratified lakes.

A Multi-Function Sensor for Eddy Correlation Measurements of Benthic Flux.

Eddy Correlation (EC) is a technique that can be used to measure transport of substances in aquatic ecosystems between bottom sediments and the overlying water (i.e. benthic fluxes). Based on high-speed, simultaneous, and co-located velocity and concentration measurements, EC has been successfully used in a variety of freshwater and marine settings to determine benthic fluxes of dissolved oxygen. Application to a larger range of compounds is limited, however, by the lack of suitable chemical sensors. Here, we describe FACT, a novel, high-speed, multi-function sensor created to expand the range of benthic fluxes that can be measured with EC. An optical fiber spectrofluorometer with a proximally located conductivity cell and thermistor, FACT enables benthic flux measurements of fluorescing compounds, such as fluorescent dissolved organic matter, as well as of heat and salinity which can be used as tracers for submarine groundwater discharge. The high bandwidth and open-beam geometry of the fluorescence sensor are particularly beneficial for EC measurements. FACT was integrated with a velocity sensor into a full EC system capable of simultaneous benthic flux measurements of fluorescing compounds, heat, and salinity. Tested in a laboratory tank, fluxes measured by all three sensors were found to track each other as well as compare favorably with expected values. Furthermore, the ability to measure fluxes of multiple substances both extends the applicability of EC to a wider range of natural sites, and can provide insight into issues of sensing volume and time responses as they affect the application of EC to natural waters.

Emerging investigator series: atmospheric cycling of indium in the northeastern United States.

Indium is critical to the global economy and is used in an increasing number of electronics and new energy technologies. However, little is known about its environmental behavior or impacts, including its concentrations or cycling in the atmosphere. This study determined indium concentrations in air particulate matter at five locations across the northeastern United States over the course of one year, in 1995. Historical records from a Massachusetts bog core showed that indium atmospheric concentrations in this region changed only modestly between 1995 and 2010. Atmospheric indium concentrations varied significantly both geographically and temporally, with average concentrations in PM3 of 2.1 ± 1.6 pg m-3 (1 standard deviation), and average particle-normalized concentrations of 0.2 ± 0.2 µg In per g PM3. Peaks in the particle-normalized concentrations in two New York sites were correlated with wind direction; air coming from the north contributed higher concentrations of indium than air coming from the west. This correlation, along with measurements of indium in zinc smelter emissions and coal fly ash, suggests that indium in the atmosphere in the northeastern United States comes from a relatively constant low-level input from coal combustion in the midwest, and higher but more sporadic contributions from the smelting of lead, zinc, copper, tin, and nickel north of the New York sample sites. Understanding the industrial sources of indium to the atmosphere and how they compare with natural sources can lead to a better understanding of the impact of human activities on the indium cycle, and may help to establish a baseline for monitoring future impacts as indium use grows.

A Prototype Sensor for In Situ Sensing of Fine Particulate Matter and Volatile Organic Compounds.

Air pollution exposure causes seven million deaths per year, according to the World Health Organization. Possessing knowledge of air quality and sources of air pollution is crucial for managing air pollution and providing early warning so that a swift counteractive response can be carried out. An optical prototype sensor (AtmOptic) capable of scattering and absorbance measurements has been developed to target in situ sensing of fine particulate matter (PM2.5) and volatile organic compounds (VOCs). For particulate matter testing, a test chamber was constructed and the emission of PM2.5 from incense burning inside the chamber was measured using the AtmOptic. The weight of PM2.5 particles was collected and measured with a filter to determine their concentration and the sensor signal-to-concentration correlation. The results of the AtmOptic were also compared and found to trend well with the Dylos DC 1100 Pro air quality monitor. The absorbance spectrum of VOCs emitted from various laboratory chemicals and household products as well as a two chemical mixtures were recorded. The quantification was demonstrated, using toluene as an example, by calibrating the AtmOptic with compressed gas standards containing VOCs at different concentrations. The results demonstrated the sensor capabilities in measuring PM2.5 and volatile organic compounds.

Methane Bubble Size Distributions, Flux, and Dissolution in a Freshwater Lake.

The majority of methane produced in many anoxic sediments is released via ebullition. These bubbles are subject to dissolution as they rise, and dissolution rates are strongly influenced by bubble size. Current understanding of natural methane bubble size distributions is limited by the difficulty in measuring bubble sizes over wide spatial or temporal scales. Our custom optical bubble size sensors recorded bubble sizes and release timing at 8 locations in Upper Mystic Lake, MA continuously for 3 months. Bubble size distributions were spatially heterogeneous even over relatively small areas experiencing similar flux, suggesting that localized sediment conditions are important to controlling bubble size. There was no change in bubble size distributions over the 3 month sampling period, but mean bubble size was positively correlated with daily ebullition flux. Bubble data was used to verify the performance of a widely used bubble dissolution model, and the model was then used to estimate that bubble dissolution accounts for approximately 10% of methane accumulated in the hypolimnion during summer stratification, and at most 15% of the diffusive air-water-methane flux from the epilimnion.

A Multi-Platform Optical Sensor for In Vivo and In Vitro Algae Classification.

Differentiation among major algal groups is important for the ecological and biogeochemical characterization of water bodies, and for practical management of water resources. It helps to discern the taxonomic groups that are beneficial to aquatic life from the organisms causing harmful algal blooms. An LED-induced fluorescence (LEDIF) instrument capable of fluorescence, absorbance, and scattering measurements; is used for in vivo and in vitro identification and quantification of four algal groups found in freshwater and marine environments. Aqueous solutions of individual and mixed dissolved biological pigments relevant to different algal groups were measured to demonstrate the LEDIF's capabilities in measuring extracted pigments. Different genera of algae were cultivated and the cell counts of the samples were quantified with a hemacytometer and/or cellometer. Dry weight of different algae cells was also measured to determine the cell counts-to-dry weight correlations. Finally, in vivo measurements of different genus of algae at different cell concentrations and mixed algal group in the presence of humic acid were performed with the LEDIF. A field sample from a local reservoir was measured with the LEDIF and the results were verified using hemacytometer, cellometer, and microscope. The results demonstrated the LEDIF's capabilities in classifying and quantifying different groups of live algae.

The precipitation of indium at elevated pH in a stream influenced by acid mine drainage.

Indium is an increasingly important metal in semiconductors and electronics and has uses in important energy technologies such as photovoltaic cells and light-emitting diodes (LEDs). One significant flux of indium to the environment is from lead, zinc, copper, and tin mining and smelting, but little is known about its aqueous behavior after it is mobilized. In this study, we use Mineral Creek, a headwater stream in southwestern Colorado severely affected by heavy metal contamination as a result of acid mine drainage, as a natural laboratory to study the aqueous behavior of indium. At the existing pH of ~3, indium concentrations are 6-29µg/L (10,000× those found in natural rivers), and are completely filterable through a 0.45µm filter. During a pH modification experiment, the pH of the system was raised to >8, and >99% of the indium became associated with the suspended solid phase (i.e. does not pass through a 0.45µm filter). To determine the mechanism of removal of indium from the filterable and likely primarily dissolved phase, we conducted laboratory experiments to determine an upper bound for a sorption constant to iron oxides, and used this, along with other published thermodynamic constants, to model the partitioning of indium in Mineral Creek. Modeling results suggest that the removal of indium from the filterable phase is consistent with precipitation of indium hydroxide from a dissolved phase. This work demonstrates that nonferrous mining processes can be a significant source of indium to the environment, and provides critical information about the aqueous behavior of indium.

Surveys, simulation and single-cell assays relate function and phylogeny in a lake ecosystem.

Much remains unknown about what drives microbial community structure and diversity. Highly structured environments might offer clues. For example, it may be possible to identify metabolically similar species as groups of organisms that correlate spatially with the geochemical processes they carry out. Here, we use a 16S ribosomal RNA gene survey in a lake that has chemical gradients across its depth to identify groups of spatially correlated but phylogenetically diverse organisms. Some groups had distributions across depth that aligned with the distributions of metabolic processes predicted by a biogeochemical model, suggesting that these groups performed biogeochemical functions. A single-cell genetic assay showed, however, that the groups associated with one biogeochemical process, sulfate reduction, contained only a few organisms that have the genes required to reduce sulfate. These results raise the possibility that some of these spatially correlated groups are consortia of phylogenetically diverse and metabolically different microbes that cooperate to carry out geochemical functions.

Statistical generation of training sets for measuring NO3(-), NH4(+) and major ions in natural waters using an ion selective electrode array.

Knowledge of ionic concentrations in natural waters is essential to understand watershed processes. Inorganic nitrogen, in the form of nitrate and ammonium ions, is a key nutrient as well as a participant in redox, acid-base, and photochemical processes of natural waters, leading to spatiotemporal patterns of ion concentrations at scales as small as meters or hours. Current options for measurement in situ are costly, relying primarily on instruments adapted from laboratory methods (e.g., colorimetric, UV absorption); free-standing and inexpensive ISE sensors for NO3(-) and NH4(+) could be attractive alternatives if interferences from other constituents were overcome. Multi-sensor arrays, coupled with appropriate non-linear signal processing, offer promise in this capacity but have not yet successfully achieved signal separation for NO3(-) and NH4(+)in situ at naturally occurring levels in unprocessed water samples. A novel signal processor, underpinned by an appropriate sensor array, is proposed that overcomes previous limitations by explicitly integrating basic chemical constraints (e.g., charge balance). This work further presents a rationalized process for the development of such in situ instrumentation for NO3(-) and NH4(+), including a statistical-modeling strategy for instrument design, training/calibration, and validation. Statistical analysis reveals that historical concentrations of major ionic constituents in natural waters across New England strongly covary and are multi-modal. This informs the design of a statistically appropriate training set, suggesting that the strong covariance of constituents across environmental samples can be exploited through appropriate signal processing mechanisms to further improve estimates of minor constituents. Two artificial neural network architectures, one expanded to incorporate knowledge of basic chemical constraints, were tested to process outputs of a multi-sensor array, trained using datasets of varying degrees of statistical representativeness to natural water samples. The accuracy of ANN results improves monotonically with the statistical representativeness of the training set (error decreases by ∼5×), while the expanded neural network architecture contributes a further factor of 2-3.5 decrease in error when trained with the most representative sample set. Results using the most statistically accurate set of training samples (which retain environmentally relevant ion concentrations but avoid the potential interference of humic acids) demonstrated accurate, unbiased quantification of nitrate and ammonium at natural environmental levels (±20% down to <10 µM), as well as the major ions Na(+), K(+), Ca(2+), Mg(2+), Cl(-), and SO4(2-), in unprocessed samples. These results show promise for the development of new in situ instrumentation for the support of scientific field work.

Atmospheric Deposition of Indium in the Northeastern United States: Flux and Historical Trends.

The metal indium is an example of an increasingly important material used in electronics and new energy technologies, whose environmental behavior and toxicity are poorly understood despite increasing evidence of detrimental health impacts and human-induced releases to the environment. In the present work, the history of indium deposition from the atmosphere is reconstructed from its depositional record in an ombrotrophic bog in Massachusetts. A novel freeze-coring technique is used to overcome coring difficulties posed by woody roots and peat compressibility, enabling retrieval of relatively undisturbed peat cores dating back more than a century. Results indicate that long-range atmospheric transport is a significant pathway for the transport of indium, with peak concentrations of 69 ppb and peak fluxes of 1.9 ng/cm2/yr. Atmospheric deposition to the bog began increasing in the late 1800s/early 1900s, and peaked in the early 1970s. A comparison of deposition data with industrial production and emissions estimates suggests that both coal combustion and the smelting of lead, zinc, copper, and tin sulfides are sources of indium to the atmosphere in this region. Deposition appears to have decreased considerably since the 1970s, potentially a visible effect of particulate emissions controls instated in North America during that decade.

Surface enhanced Raman scattering by graphene-nanosheet-gapped plasmonic nanoparticle arrays for multiplexed DNA detection.

We have developed a new type of surface enhanced Raman scattering (SERS) substrate with thiolated graphene oxide (tGO) nanosheets sandwiched between two layers of closely packed plasmonic nanoparticles. The trilayered substrate is built up through alternative loading of interfacially assembled plasmonic nanoparticle arrays and tGO nanosheets, followed by coating the nanoparticle surfaces with poly(ethylene glycol) (PEG). Here tGO plays multifunctional roles as a 2D scaffold to immobilized interfacially assembled plasmonic nanoparticles, a nanospacer to create SERS-active nanogaps between two layers of nanoparticle arrays, and a molecule harvester to enrich molecules of interest viaπ-π interaction. In particular, the molecule harvesting capability of the tGO nanospacer and the stealth properties of PEG coating on the plasmonic nanoparticles collectively lead to preferential positioning of selective targets such as aromatic molecules and single-stranded DNA at the SERS-active nanogap hotspots. We have demonstrated that an SERS assay based on the PEGylated trilayered substrate, in combination with magnetic separation, allows for sensitive, multiplexed "signal-off" detection of DNA sequences of bacterial pathogens.

Methane cycling. Nonequilibrium clumped isotope signals in microbial methane.

Methane is a key component in the global carbon cycle, with a wide range of anthropogenic and natural sources. Although isotopic compositions of methane have traditionally aided source identification, the abundance of its multiply substituted "clumped" isotopologues (for example, (13)CH3D) has recently emerged as a proxy for determining methane-formation temperatures. However, the effect of biological processes on methane's clumped isotopologue signature is poorly constrained. We show that methanogenesis proceeding at relatively high rates in cattle, surface environments, and laboratory cultures exerts kinetic control on (13)CH3D abundances and results in anomalously elevated formation-temperature estimates. We demonstrate quantitatively that H2 availability accounts for this effect. Clumped methane thermometry can therefore provide constraints on the generation of methane in diverse settings, including continental serpentinization sites and ancient, deep groundwaters.

Extended artificial neural networks: incorporation of a priori chemical knowledge enables use of ion selective electrodes for in-situ measurement of ions at environmentally relevant levels.

A novel artificial neural network (ANN) architecture is proposed which explicitly incorporates a priori system knowledge, i.e., relationships between output signals, while preserving the unconstrained non-linear function estimator characteristics of the traditional ANN. A method is provided for architecture layout, disabling training on a subset of neurons, and encoding system knowledge into the neuron structure. The novel architecture is applied to raw readings from a chemical sensor multi-probe (electric tongue), comprised of off-the-shelf ion selective electrodes (ISEs), to estimate individual ion concentrations in solutions at environmentally relevant concentrations and containing environmentally representative ion mixtures. Conductivity measurements and the concept of charge balance are incorporated into the ANN structure, resulting in (1) removal of estimation bias typically seen with use of ISEs in mixtures of unknown composition and (2) improvement of signal estimation by an order of magnitude or more for both major and minor constituents relative to use of ISEs as stand-alone sensors and error reduction by 30-50% relative to use of standard ANN models. This method is suggested as an alternative to parameterization of traditional models (e.g., Nikolsky-Eisenman), for which parameters are strongly dependent on both analyte concentration and temperature, and to standard ANN models which have no mechanism for incorporation of system knowledge. Network architecture and weighting are presented for the base case where the dot product can be used to relate ion concentrations to both conductivity and charge balance as well as for an extension to log-normalized data where the model can no longer be represented in this manner. While parameterization in this case study is analyte-dependent, the architecture is generalizable, allowing application of this method to other environmental problems for which mathematical constraints can be explicitly stated.

Towards an automated, standardized protocol for determination of equilibrium potential of ion-selective electrodes.

An automated real-time method for determination of ISE steady state value and response time is developed, following most recent IUPAC recommendations. Specifically, detection of the 'steady state' is related to (1) the time derivative of the emf as it reaches a limiting value (ΔE/Δt(limit), e.g., 0.1-1.0 mV min(-1)) and (2) the duration of time for which the absolute value of the time derivative remains less than this limiting value (stability window, denoted win(st)). A suite of representative ISEs, including glass, solid state, and polymer-based electrodes, is examined to determine sensitivity of results to parameterization choice. Measurements taken over a wide range of concentration values and in un-processed samples (i.e., without use of ionic strength adjustment) provide insight into behavior of ISEs in applications where analyte concentrations span a wide range and/or sample pre-processing may not be an option, e.g., use of sensors for in situ environmental sampling. Results show that declared steady state emf is strongly sensitive to variations in ΔE/Δt(limit) but relatively unaffected by changes in the stability window when win(st) ≥30 s. Linearity of calibration curves produced, quantified by root mean squared error (RMSE) against a linear fit, improves as ΔE/Δt(limit) decreases, however the percentage of measurements which reach a declared steady state within the prescribed sample window (∼6.5 min) falls with corresponding decreases in the ΔE/Δt(limit) parameter. Response time, defined as the time required to reach declared steady emf, is also a strong function of parameterization. Dependence of response times on sample composition and/or ISE membrane composition and type are also discussed; results for ISEs in samples comprised exclusively of interfering ions are included. In general, limiting emf derivatives of {0.25-0.4 mV min(-1)} and stability windows of {30-40s} achieve both good analytical accuracy and compliance with potentially short sampling window requirements. Methodology based on use of these parameters can improve sampling speed and accuracy as well as promote inter-comparison of data and ISE characterizations among research teams.

Nitrate suppresses internal phosphorus loading in an eutrophic lake.

The presence of nitrate in the hypolimnion of the eutrophic, dimictic Upper Mystic Lake has been previously shown to suppress the release of arsenic from lake sediments during seasonal anoxia, in large part by oxidizing iron (II) and producing iron oxyhydroxides that sorb inorganic arsenic. Because of the importance of internal phosphorus loading in the phosphorus budget of many eutrophic lakes, the chemical similarities between phosphate and arsenate, and the need to account for internal phosphorus loading as part of many lake restoration strategies, we carried out measurements to determine if the presence of nitrate also suppressed the release of phosphorus from the sediments of this lake during anoxia. Observations showed that this was the case. Arsenic, phosphorus, and iron (II) concentrations were strongly correlated in the water column, as expected, and the depths below which phosphorus and iron concentrations increased relative to epilimnetic values was predicted by the depth at which nitrate concentration approached zero. The results suggest that knowledge of a lake's nitrogen budget may be a useful tool in the design of lake remediation efforts, even though phosphorus is typically the limiting nutrient.

Anthropogenic forcings on the surficial osmium cycle.

Osmium is among the least abundant elements in the Earth's continental crust. Recent anthropogenic Os contamination of the environment from mining and smelting activities, automotive catalytic converter use, and hospital discharges has been documented. Here we present evidence for anthropogenic overprinting of the natural Os cycle using a ca. 7000-year record of atmospheric Os deposition and isotopic composition from an ombrotrophic peat bog in NW Spain. Preanthropogenic Os accumulation in this area is 0.10 +/- 0.04 ng m(-2) y(-1). The oldest strata showing human influence correspond to early metal mining and processing on the Iberian Peninsula (ca. 4700-2500 cal. BP). Elevated Os accumulation rates are found thereafter with a local maximum of 1.1 ng m(-2) y(-1) during the Roman occupation of the Iberian Peninsula (ca. 1930 cal. BP) and a further increase starting in 1750 AD with Os accumulation reaching 30 ng m(-2) y(-1) in the most recent samples. Osmium isotopic composition ((187)Os/(188)Os) indicates that recent elevated Os accumulation results from increased input of unradiogenic Os from industrial and automotive sources as well as from enhanced deposition of radiogenic Os through increased fossil fuel combustion and soil erosion. We posit that the rapid increase in catalyst-equipped vehicles, increased fossil fuel combustion, and changes in land-use make the changes observed in NW Spain globally relevant.

A Differential Pressure Instrument with Wireless Telemetry for In-Situ Measurement of Fluid Flow across Sediment-Water Boundaries.

An instrument has been built to carry out continuous in-situ measurement of small differences in water pressure, conductivity and temperature, in natural surface water and groundwater systems. A low-cost data telemetry system provides data on shore in real time if desired. The immediate purpose of measurements by this device is to continuously infer fluxes of water across the sediment-water interface in a complex estuarine system; however, direct application to assessment of sediment-water fluxes in rivers, lakes, and other systems is also possible. Key objectives of the design include both low cost, and accuracy of the order of ±0.5 mm H(2)O in measured head difference between the instrument's two pressure ports. These objectives have been met, although a revision to the design of one component was found to be necessary. Deployments of up to nine months, and wireless range in excess of 300 m have been demonstrated.

Phylogenetic analysis implicates birds as a source of Cryptosporidium spp. oocysts in agricultural watersheds.

Cryptosporidium parvum and C. hominis are protozoan parasites responsible for cryptosporidiosis, an acute gastrointestinal illness that can be life-threatening for immunocompromised persons. Sources and genotypes of Cryptosporidium oocysts were investigated in two agricultural areas within the Wachusett Reservoir watershed, a drinking water source for Boston, Massachusetts. Two brooks (denoted Brook SF and Brook JF, respectively), each downgradient from a dairy farm, were chosen as sample sites. For one year, Brooks SF and JF were sampled monthly; oocysts were detected in 6 (50%) out of 12 samples from Brook JF, and no oocysts were detected in Brook SF. Oocyst genotypes from agricultural surface waters were compared to oocyst genotypes from Genbank, as well as fecal samples of cattle and birds, using phylogenetic analysis of a hypervariable region of the 18S rRNA gene by both neighbor-joining and parsimony methods. Results show extensive heterogeneity among Cryptosporidium spp. 18S rRNA sequences, and also suggest that birds are an oocyst source in this watershed. Principal components analysis showed oocyst presence correlating strongly with seasonal factors, and oocysts in surface waters were only detected in the summer through late fall, co-incident with the presence of migratory birds in this watershed. If birds are confirmed to be an important source of oocysts infectious to humans, the data suggest that protection of raw drinking water supplies in some agricultural areas may depend upon management and control of resident and migratory bird populations.