Critical considerations for communicating environmental DNA science.

The economic and methodological efficiencies of environmental DNA (eDNA) based survey approaches provide an unprecedented opportunity to assess and monitor aquatic environments. However, instances of inadequate communication from the scientific community about confidence levels, knowledge gaps, reliability, and appropriate parameters of eDNA-based methods have hindered their uptake in environmental monitoring programs and, in some cases, has created misperceptions or doubts in the management community. To help remedy this situation, scientists convened a session at the Second National Marine eDNA Workshop to discuss strategies for improving communications with managers. These include articulating the readiness of different eDNA applications, highlighting the strengths and limitations of eDNA tools for various applications or use cases, communicating uncertainties associated with specified uses transparently, and avoiding the exaggeration of exploratory and preliminary findings. Several key messages regarding implementation, limitations, and relationship to existing methods were prioritized. To be inclusive of the diverse managers, practitioners, and researchers, we and the other workshop participants propose the development of communication workflow plans, using RACI (Responsible, Accountable, Consulted, Informed) charts to clarify the roles of all pertinent individuals and parties and to minimize the chance for miscommunications. We also propose developing decision support tools such as Structured Decision-Making (SDM) to help balance the benefits of eDNA sampling with the inherent uncertainty, and developing an eDNA readiness scale to articulate the technological readiness of eDNA approaches for specific applications. These strategies will increase clarity and consistency regarding our understanding of the utility of eDNA-based methods, improve transparency, foster a common vision for confidently applying eDNA approaches, and enhance their benefit to the monitoring and assessment community.

Toward improved sediment management and coastal resilience through efficient permitting in California.

The value of sediment for helping coastal habitats and infrastructure respond to sea level rise is widely recognized. Across the country, coastal managers are seeking ways to beneficially use sediment sourced from dredging and other projects to counter coastal erosion and protect coastal resources. However, these projects are difficult to permit and have been slow to actualize. This paper draws on interviews with sediment managers and regulators in California to explore the challenges and opportunities for habitat restoration and beach nourishment within the current permitting regime. We find that permits are costly, difficult to obtain, and sometimes stand as a barrier to more sustainable and adaptive sediment management. We next characterize streamlining approaches and describe entities and ongoing efforts within California that apply them. Finally, we conclude that to keep pace with coastal losses due to climate change impacts, efforts toward efficient permitting must be accelerated and approaches diversified to support coastal resilience practices state-wide, in a timeframe that will allow coastal managers to innovate and adapt.

Simulating the thermal impact of substrate temperature on ecological restoration in shallow urban rivers.

Managing river temperature in highly urbanized stream systems is critical for maintaining aquatic ecosystems and associated beneficial uses. In this work, we updated and utilized a mechanistic river temperature model, i-Tree Cool River, to evaluate the cooling impacts of two ecological restoration scenarios: (1) an alternative streambed material limecrete and (2) shading effects of tree planting in riparian areas. The i-Tree Cool River model was modified to account for diurnal fluctuations of streambed temperature, which is relevant in shallow urban streams where lack of natural shading combined with low heat capacity of the water column can make diurnal fluctuations relatively extreme. The model was calibrated and validated on a 4.2 km reach of Compton Creek in the Los Angeles River watershed, California. Two native fish, arroyo chub (Gila orcuttii) and unarmored threespine stickleback (Gasterosteus aculeatus williamsoni), were considered the target species for assessing thermal habitat suitability. Key findings include: (1) model performance was improved when accounting for diurnal fluctuations in bed temperature (R2 increased from 0.43 to 0.68); and (2) substrate rehabilitation and tree planting can potentially reduce summertime temperatures to within the documented spawning temperature thresholds for the focal fish species. Using limecrete as an alternative material for the concrete bottom decreased the median river temperature metrics: maximum weekly maximum, maximum weekly average, and minimum weekly minimum temperatures by an average of 3 °C (13%) to 20.4 °C, 19.7 °C, and 17.8 °C, respectively. Tree planting in the riparian corridor decreased the average river temperature metrics by an average of 0.9 °C (4%) to 22.7 °C, 22 °C, and 19 °C, respectively. Combining the two scenarios decreased the river temperature metrics by an average of 4 °C (18%) to 18.2 °C. Therefore, water temperature would not be a limiting factor in potential reintroduction of the focal fish species to Compton Creek if restoration were implemented. Implications of this work could be used by urban forest and water managers for restoring thermally polluted rivers in other urban areas.

The impact of climate change induced alterations of streamflow and stream temperature on the distribution of riparian species.

Distributions of riparian species will likely shift due to climate change induced alterations in temperature and rainfall patterns, which alter stream habitat. Spatial forecasting of suitable habitat in projected climatic conditions will inform management interventions that support wildlife. Challenges in developing forecasts include the need to consider the large number of riparian species that might respond differently to changing conditions and the need to evaluate the many different characteristics of streamflow and stream temperature that drive species-specific habitat suitability. In particular, in dynamic environments like streams, the short-term temporal resolution of species occurrence and streamflow need to be considered to identify the types of conditions that support various species. To address these challenges, we cluster species based on habitat characteristics to select habitat representatives and we evaluate regional changes in habitat suitability using short-term, temporally explicit metrics that describe the streamflow and stream temperature regime. We use stream-specific environmental predictors rather than climatic variables. Unlike other studies, the stream-specific environmental predictors are generated from the time that species were observed in a particular reach, in addition to long term trends, to evaluate habitat preferences. With species occurrence data from local monitoring surveys and streamflow and stream temperature modeled from downscaled Coupled Model Intercomparison Project - Phase 5 (CMIP5) climate projections, we predict change in habitat suitability at the end-of-century. The relative importance of hydrology and stream temperature varied by cluster. High altitudinal, cold water species' distributions contracted, while lower elevation, warm water species distributions expanded. Modeling with short-term temporally explicit environmental metrics did produce different end-of-century projections than using long-term averages for some of the representative species. These findings can help wildlife managers prioritize conservation efforts, manage streamflow, initiate monitoring of species in vulnerable clusters, and address stressors, such as passage barriers, in areas projected to be suitable in future climate conditions.

Translational Molecular Ecology in practice: Linking DNA-based methods to actionable marine environmental management.

Molecular-based approaches can provide timely biodiversity assessments, showing an immense potential to facilitate decision-making in marine environmental management. However, the uptake of molecular data into environmental policy remains minimal. Here, we showcase a selection of local to global scale studies applying molecular-based methodologies for environmental management at various stages of implementation. Drawing upon lessons learned from these case-studies, we provide a roadmap to facilitate applications of DNA-based methods to marine policies and to overcome the existing challenges. The main impediment identified is the need for standardized protocols to guarantee data comparison across spatial and temporal scales. Adoption of Translational Molecular Ecology - the sustained collaboration between molecular ecologists and stakeholders, will enhance consensus with regards to the objectives, methods, and outcomes of environmental management projects. Establishing a sustained dialogue among stakeholders is key to accelerating the adoption of molecular-based approaches for marine monitoring and assessment.

Using alpha, beta, and zeta diversity in describing the health of stream-based benthic macroinvertebrate communities.

Ecological monitoring of streams has frequently focused on measures describing the taxonomic, and sometimes functional, α diversity of benthic macroinvertebrates (BMIs) within a single sampled community. However, as many ecological processes effectively link BMI stream communities there is a need to describe groups of communities using measures of regional diversity. Here we demonstrate a role for incorporating both a traditional pairwise measure of community turnover, ß diversity, in assessing community health as well as ζ diversity, a more generalized framework for describing similarity between multiple communities. Using 4,395 samples of BMI stream communities in California, we constructed a model using measures of α, ß, and ζ diversity, which accounted for 71.7% of among-watershed variation in the mean health of communities, as described by the California Streams Condition Index (CSCI). We also investigated the use of ζ diversity in assessing models of stochastic vs. niche assembly across communities of BMIs within watersheds, with the niche assembly model found to be the likelier of the two.

Evaluating regional resiliency of coastal wetlands to sea level rise through hypsometry-based modeling.

Sea level rise (SLR) threatens coastal wetlands worldwide, yet the fate of individual wetlands will vary based on local topography, wetland morphology, sediment dynamics, hydrologic processes, and plant-mediated feedbacks. Local variability in these factors makes it difficult to predict SLR effects across wetlands or to develop a holistic regional perspective on SLR response for a diversity of wetland types. To improve regional predictions of SLR impacts to coastal wetlands, we developed a model that addresses the scale-dependent factors controlling SLR response and accommodates different levels of data availability. The model quantifies SLR-driven habitat conversion within wetlands across a region by predicting changes in individual wetland hypsometry. This standardized approach can be applied to all wetlands in a region regardless of data availability, making it ideal for modeling SLR response across a range of scales. Our model was applied to 105 wetlands in southern California that spanned a broad range of typology and data availability. Our findings suggest that if wetlands are confined to their current extents, the region will lose 12% of marsh habitats (vegetated marsh and unvegetated flats) with 0.6 m of SLR (projected for 2050) and 48% with 1.7 m of SLR (projected for 2100). Habitat conversion was more drastic in wetlands with larger proportions of marsh habitats relative to subtidal habitats and occurred more rapidly in small lagoons relative to larger sites. Our assessment can inform management of coastal wetland vulnerability, improve understanding of the SLR drivers relevant to individual wetlands, and highlight significant data gaps that impede SLR response modeling across spatial scales. This approach augments regional SLR assessments by considering spatial variability in SLR response drivers, addressing data gaps, and accommodating wetland diversity, which will provide greater insights into regional SLR response that are relevant to coastal management and restoration efforts.

Beyond Metrics? The Role of Hydrologic Baseline Archetypes in Environmental Water Management.

Balancing ecological and human water needs often requires characterizing key aspects of the natural flow regime and then predicting ecological response to flow alterations. Flow metrics are generally relied upon to characterize long-term average statistical properties of the natural flow regime (hydrologic baseline conditions). However, some key aspects of hydrologic baseline conditions may be better understood through more complete consideration of continuous patterns of daily, seasonal, and inter-annual variability than through summary metrics. Here we propose the additional use of high-resolution dimensionless archetypes of regional stream classes to improve understanding of baseline hydrologic conditions and inform regional environmental flows assessments. In an application to California, we describe the development and analysis of hydrologic baseline archetypes to characterize patterns of flow variability within and between stream classes. We then assess the utility of archetypes to provide context for common flow metrics and improve understanding of linkages between aquatic patterns and processes and their hydrologic controls. Results indicate that these archetypes may offer a distinct and complementary tool for researching mechanistic flow-ecology relationships, assessing regional patterns for streamflow management, or understanding impacts of changing climate.

Microcystin Prevalence throughout Lentic Waterbodies in Coastal Southern California.

Toxin producing cyanobacterial blooms have increased globally in recent decades in both frequency and intensity. Despite the recognition of this growing risk, the extent and magnitude of cyanobacterial blooms and cyanotoxin prevalence is poorly characterized in the heavily populated region of southern California. Recent assessments of lentic waterbodies (depressional wetlands, lakes, reservoirs and coastal lagoons) determined the prevalence of microcystins and, in some cases, additional cyanotoxins. Microcystins were present in all waterbody types surveyed although toxin concentrations were generally low across most habitats, as only a small number of sites exceeded California's recreational health thresholds for acute toxicity. Results from passive samplers (Solid Phase Adsorption Toxin Tracking (SPATT)) indicated microcystins were prevalent throughout lentic waterbodies and that traditional discrete samples underestimated the presence of microcystins. Multiple cyanotoxins were detected simultaneously in some systems, indicating multiple stressors, the risk of which is uncertain since health thresholds are based on exposures to single toxins. Anatoxin-a was detected for the first time from lakes in southern California. The persistence of detectable microcystins across years and seasons indicates a low-level, chronic risk through both direct and indirect exposure. The influence of toxic cyanobacterial blooms is a more complex stressor than presently recognized and should be included in water quality monitoring programs.

Is DNA barcoding actually cheaper and faster than traditional morphological methods: results from a survey of freshwater bioassessment efforts in the United States?

Taxonomic identification accounts for a substantial portion of cost associated with bioassessment programs across the United States. New analytical approaches, such as DNA barcoding have been promoted as a way to reduce monitoring costs and improve efficiency, yet this assumption has not been thoroughly evaluated. We address this question by comparing costs for traditional morphology-based bioassessment, the standard Sanger sequencing-based DNA barcoding approach, and emerging next-generation (NGS) molecular methods. Market demand for molecular approaches is also assessed through a survey of the level of freshwater bioassessment effort in the United States across multiple habitat types (lakes, streams, wetlands) and indicators (benthic invertebrates, fish, algae). All state and regional level programs administered by public agencies and reported via agency web sites were included in the survey. Costs were based on surveys of labs and programs willing to provide such information. More than 19,500 sites are sampled annually across the United States, with the majority of effort occurring in streams. Benthic invertebrates are the most commonly used indicator, but algae and fish comprise between 35% and 21% of total sampling effort, respectively. We estimate that between $104 and $193 million is spent annually on routine freshwater bioassessment in the United States. Approximately 30% of the bioassessment costs are comprised of the cost to conduct traditional morphology-based taxonomy. Current barcoding costs using Sanger sequencing are between 1.7 and 3.4 times as expensive as traditional taxonomic approaches, excluding the cost of field sampling (which is common to both approaches). However, the cost of NGS methods are comparable (or slightly less expensive) than traditional methods depending on the indicator. The promise of barcoding as a cheaper alternative to current practices is not yet realized, although molecular methods may provide other benefits, such as a faster sample processing and increased taxonomic resolution.

Selecting the optimum plot size for a California design-based stream and wetland mapping program.

Accurate estimates of the extent and distribution of wetlands and streams are the foundation of wetland monitoring, management, restoration, and regulatory programs. Traditionally, these estimates have relied on comprehensive mapping. However, this approach is prohibitively resource-intensive over large areas, making it both impractical and statistically unreliable. Probabilistic (design-based) approaches to evaluating status and trends provide a more cost-effective alternative because, compared with comprehensive mapping, overall extent is inferred from mapping a statistically representative, randomly selected subset of the target area. In this type of design, the size of sample plots has a significant impact on program costs and on statistical precision and accuracy; however, no consensus exists on the appropriate plot size for remote monitoring of stream and wetland extent. This study utilized simulated sampling to assess the performance of four plot sizes (1, 4, 9, and 16 km(2)) for three geographic regions of California. Simulation results showed smaller plot sizes (1 and 4 km(2)) were most efficient for achieving desired levels of statistical accuracy and precision. However, larger plot sizes were more likely to contain rare and spatially limited wetland subtypes. Balancing these considerations led to selection of 4 km(2) for the California status and trends program.

Evaluating ethanol-based sample preservation to facilitate use of DNA barcoding in routine freshwater biomonitoring programs using benthic macroinvertebrates.

Molecular methods, such as DNA barcoding, have the potential to enhance biomonitoring programs worldwide. Altering routinely used sample preservation methods to protect DNA from degradation may pose a potential impediment to application of DNA barcoding and metagenomics for biomonitoring using benthic macroinvertebrates. Using higher volumes or concentrations of ethanol, requirements for shorter holding times, or the need to include additional filtering may increase cost and logistical constraints to existing biomonitoring programs. To address this issue we evaluated the efficacy of various ethanol-based sample preservation methods at maintaining DNA integrity. We evaluated a series of methods that were minimally modified from typical field protocols in order to identify an approach that can be readily incorporated into existing monitoring programs. Benthic macroinvertebrates were collected from a minimally disturbed stream in southern California, USA and subjected to one of six preservation treatments. Ten individuals from five taxa were selected from each treatment and processed to produce DNA barcodes from the mitochondrial gene cytochrome c oxidase I (COI). On average, we obtained successful COI sequences (i.e. either full or partial barcodes) for between 93-99% of all specimens across all six treatments. As long as samples were initially preserved in 95% ethanol, successful sequencing of COI barcodes was not affected by a low dilution ratio of 2∶1, transfer to 70% ethanol, presence of abundant organic matter, or holding times of up to six months. Barcoding success varied by taxa, with Leptohyphidae (Ephemeroptera) producing the lowest barcode success rate, most likely due to poor PCR primer efficiency. Differential barcoding success rates have the potential to introduce spurious results. However, routine preservation methods can largely be used without adverse effects on DNA integrity.

Metals and bacteria partitioning to various size particles in Ballona Creek storm water runoff.

Many storm water best management practice (BMP) devices function primarily by capturing particulate matter to take advantage of the well-documented association between storm water particles and pollutants. The hydrodynamic separation or settling methods used by most BMP devices are most effective at capturing medium to large particles; however, these may not be the most predominant particles associated with urban runoff. The present study examined particle size distribution in storm water runoff from an urban watershed in southern California and investigated the pollutant-particle associations of metals (Cu, Pb, Ni, and Zn) and bacteria (enterococci and Escherichia coli). During small storm events (≤0.7 cm rain), the highest concentration of pollutants were associated with a <6-µm filter fraction, which accounted for 70% of the per storm contaminant mass but made up more than 20% of the total particle mass. The pollutant-particle association changed with storm size. Most pollutant mass was associated with >35 µm size particles during a 5-cm rain event. These results suggest that much of the contaminant load in storm water runoff will not be captured by the most commonly used BMP devices, because most of these devices (e.g., hydrodynamic separators) are unable to capture particles smaller than 75 µm.

Stormwater contaminant loading following southern California wildfires.

Contaminant loading associated with stormwater runoff from recently burned areas is poorly understood, despite the fact that it has the potential to affect downstream water quality. The goal of the present study is to assess regional patterns of runoff and contaminant loading from wildfires in urban fringe areas of southern California. Postfire stormwater runoff was sampled from five wildfires that each burned between 115 and 658 km(2) of natural open space between 2003 and 2009. Between two and five storm events were sampled per site over the first one to two years following the fires for basic constituents, metals, nutrients, total suspended solids, and polycyclic aromatic hydrocarbons (PAHs). Results were compared to data from 16 unburned natural areas and six developed sites. Mean copper, lead, and zinc flux (kg/km(2)) were between 112- and 736-fold higher from burned catchments and total phosphorus was up to 921-fold higher compared to unburned natural areas. Polycyclic aromatic hydrocarbon flux was four times greater from burned areas than from adjacent urban areas. Ash fallout on nearby unburned watersheds also resulted in a threefold increase in metals and PAHs. Attenuation of elevated concentration and flux values appears to be driven mainly by rainfall magnitude. Contaminant loading from burned landscapes has the potential to be a substantial contribution to the total annual load to downstream areas in the first several years following fires.

Performance of two Southern California benthic community condition indices using species abundance and presence-only data: relevance to DNA barcoding.

DNA barcoding, as it is currently employed, enhances use of marine benthic macrofauna as environmental condition indicators by improving the speed and accuracy of the underlying taxonomic identifications. The next generation of barcoding applications, processing bulk environmental samples, will likely only provide presence information. However, macrofauna indices presently used to interpret these data are based on species abundances. To assess the importance of this difference, we evaluated the performance of the Southern California Benthic Response Index (BRI) and the AZTI Marine Biotic Index (AMBI) when species abundance data were removed from their calculation. Presence only versions of these two indices were created by eliminating abundance weighting while preserving species identity. Associations between the presence and abundance BRI, and the presence and abundance AMBI were highly significant, with correlation coefficients of 0.99 and 0.81, respectively. The presence versions validated almost equally to the abundance-based indices when applied to the spatial and the temporal monitoring data used to validate the original indices. Simulations in which taxa were systematically removed from calculation of the indices were also conducted to assess how large the barcode library must be for the indices to be effective. Correlation between the BRI-P and BRI remained above 0.9 with only 370 species in the library and reducing the number of species to 450 had almost no effect on correlation between the presence and abundance versions of the AMBI.

Levels and patterns of fecal indicator bacteria in stormwater runoff from homogenous land use sites and urban watersheds.

Routine stormwater monitoring programs focus on quantification of average fecal indicator bacteria (FIB) concentration at the terminal watershed discharge point. While important for permit compliance, such monitoring provides little insight into relative bacteria levels from different land use types or the mechanisms that influence FIB concentrations. The goal of this study was to quantify the relative levels and flux patterns of Escherichia coli, enterococci, and total coliforms from representative land use (LU) types. Bacteria concentrations were measured over the entire storm duration from 8 different LU types over 13 storm events in 5 southern California watersheds during the 2000-2005 storm seasons. In addition, runoff samples were collected from 8 bottom of the watershed mass emission (ME) sites. Intra-storm and intra-season patterns were investigated in order to identify mechanisms that influence patterns of FIB concentrations. Mean FIB event mean concentrations (EMCs) at LU sites ranged from 10(3) to 10(5) MPN/100 mi. Recreational (horse stables) LU sites contributed significantly higher storm EMCs than other LU types. Early season storms repeatedly produced higher EMCs than comparably sized late season storms. For most storms sampled, the highest bacterial concentrations occurred during the early phases of stormwater runoff with peak concentrations usually preceding peak flow.

Evaluating performance of stormwater sampling approaches using a dynamic watershed model.

Accurate quantification of stormwater pollutant levels is essential for estimating overall contaminant discharge to receiving waters. Numerous sampling approaches exist that attempt to balance accuracy against the costs associated with the sampling method. This study employs a novel and practical approach of evaluating the accuracy of different stormwater monitoring methodologies using stormflows and constituent concentrations produced by a fully validated continuous simulation watershed model. A major advantage of using a watershed model to simulate pollutant concentrations is that a large number of storms representing a broad range of conditions can be applied in testing the various sampling approaches. Seventy-eight distinct methodologies were evaluated by "virtual samplings" of 166 simulated storms of varying size, intensity and duration, representing 14 years of storms in Ballona Creek near Los Angeles, California. The 78 methods can be grouped into four general strategies: volume-paced compositing, time-paced compositing, pollutograph sampling, and microsampling. The performances of each sampling strategy was evaluated by comparing the (1) median relative error between the virtually sampled and the true modeled event mean concentration (EMC) of each storm (accuracy), (2) median absolute deviation about the median or "MAD" of the relative error or (precision), and (3) the percentage of storms where sampling methods were within 10% of the true EMC (combined measures of accuracy and precision). Finally, costs associated with site setup, sampling, and laboratory analysis were estimated for each method. Pollutograph sampling consistently outperformed the other three methods both in terms of accuracy and precision, but was the most costly method evaluated. Time-paced sampling consistently underestimated while volume-paced sampling over estimated the storm EMCs. Microsampling performance approached that of pollutograph sampling at a substantial cost savings. The most efficient method for routine stormwater monitoring in terms of a balance between performance and cost was volume-paced microsampling, with variable sample pacing to ensure that the entirety of the storm was captured. Pollutograph sampling is recommended if the data are to be used for detailed analysis of runoff dynamics.

Ecosystem response to regulatory and management actions: the southern California experience in long-term monitoring.

Billions of dollars have been invested over the past 35 years in reducing pollutant emissions to coastal environments. Evaluation of the effectiveness of this investment is hampered by the lack of long-term consistent data. A rare opportunity exists in southern California to evaluate the effectiveness of management actions by analyzing long-term monitoring of effluent, sediment, benthos, and fish and comparing this trend data to periodic regional surveys of environmental condition. In this paper, we ask the question "have improvements in effluent quality in response to environmental regulation translated into improvements in the receiving environment?" Results indicate that management actions directed at reducing mass emissions from wastewater treatment plants (POTWs) have resulted in substantial improvement in aquatic communities. However, the magnitude and timing of response varies by indicator suggesting that use of multiple assessment endpoints is necessary to adequately interpret trends. Reductions in the effect of POTW effluent have allowed managers to shift resources to address other contaminant sources such as stormwater and resuspension of legacy pollutants.

How effective has the Clean Water Act been at reducing pollutant mass emissions to the Southern California Bight over the past 35 years?

The Clean Water Act (CWA) has regulated discharges of contaminants since 1972. However, evaluations of the CWA's effectiveness at improving regional water quality are lacking, primarily because integration of monitoring data from multiple dischargers to assess cumulative effects is not required. A rare opportunity exists to assess CWA effectiveness by integrating mass emissions data from all major sources of contaminants to the Southern California Bight from 1971 to 2000. While the coastal population grew by 56% and total effluent volume increased 31% since 1971, mass emissions of nearly all constituents decreased since passage of the CWA, most by greater than 65%. Publicly owned treatment works were the dominant point source of many contaminants, but also accounted for the greatest reductions in pollutant discharge since 1971. As point source treatment has improved, the relative contribution of non-point sources, such as storm water runoff has increased. Despite the increased importance of storm water discharges, regional monitoring and data compilation of this source is lacking, making it difficult to accurately assess trends in non-point source discharge.

Fecal indicator bacteria (FIB) levels during dry weather from Southern California reference streams.

High levels of fecal indicator bacteria (FIB) in surface waters is a common problem in urban areas that often leads to impairment of beneficial uses such as swimming. Once impaired, common management and regulatory solutions include development of total maximum daily loads (TMDLs) and other water quality management plans. A critical element of these plans is establishment of a "reference" level of exceedances against which to assess management goals and TMDL compliance. The goal of this study was to provide information on indicator bacteria contributions from natural streams in undeveloped catchments throughout southern California during dry weather, non-storm conditions. To help establish a regional reference data set, bacteria levels [i.e. Escherichia coli (E. coli), enterococci and total coliforms] were measured from 15 unimpaired streams in 11 southern California watersheds weekly for one full year. Concentrations measured from reference areas were typically between one to two orders of magnitude lower than levels found in developed watersheds. Nearly 82% of the time, samples did not exceed daily and monthly bacterial indicator thresholds. E. coli had the lowest daily percent exceedance (1.5%). A total of 13.7% of enterococci exceeded daily thresholds. Indicator bacteria levels fluctuated seasonally with an average of 79% of both enterococci and total coliforms exceedances occurring during summer months (June to August). Temperature, at all sites, explained about one-half the variation in total coliforms density suggesting that stream temperatures regulated bacterial populations. Accounting for natural background levels will allow for management targets that are more reflective of the contributions from natural sources.