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USGS2020

Shifts in hatching date of American crocodile (Crocodylus acutus) in southern Florida

Globally temperature of marine environments is on the rise and temperature plays an important role in the life-history of reptiles. In this study, we examined the relationship between sea surface temperature and average date of hatching for American crocodiles (Crocodylus acutus) over a 37-year period at two nesting sites, Everglades National Park and Florida Power and Light Turkey Point Power Plant site in southern Florida. Our results indicate that hatch dates are shifting 1.5 days earlier every two years and at half that rate for the Turkey Point site, and with every 1 °C degree increase in temperature, hatching occurs about 10 days earlier in the Everglades and 6 days earlier at Turkey Point. Our results on shifting hatch dates for American crocodiles provide further details about the impacts of temperature change on crocodile life history and suggest that increased temperature may affect their phenology.

Florida↗

USGS2015

Stock-specific advection of larval walleye ( Sander vitreus ) in western Lake Erie: Implications for larval growth, mixing, and stock discrimination

Physical processes can generate spatiotemporal heterogeneity in habitat quality for fish and also influence the overlap of pre-recruit individuals (e.g., larvae) with high-quality habitat through hydrodynamic advection. In turn, individuals from different stocks that are produced in different spawning locations or at different times may experience dissimilar habitat conditions, which can underlie within- and among-stock variability in larval growth and survival. While such physically-mediated variation has been shown to be important in driving intra- and inter-annual patterns in recruitment in marine ecosystems, its role in governing larval advection, growth, survival, and recruitment has received less attention in large lake ecosystems such as the Laurentian Great Lakes. Herein, we used a hydrodynamic model linked to a larval walleye ( Sander vitreus ) individual-based model to explore how the timing and location of larval walleye emergence from several spawning sites in western Lake Erie (Maumee, Sandusky, and Detroit rivers; Ohio reef complex) can influence advection pathways and mixing among these local spawning populations (stocks), and how spatiotemporal variation in thermal habitat can influence stock-specific larval growth. While basin-wide advection patterns were fairly similar during 2011 and 2012, smaller scale advection patterns and the degree of stock mixing varied both within and between years. Additionally, differences in larval growth were evident among stocks and among cohorts within stocks which were attributed to spatiotemporal differences in water temperature. Using these findings, we discuss the value of linked physical–biological models for understanding the recruitment process and addressing fisheries management problems in the world's Great Lakes.

Journal of Great Lakes Research↗

USGS2001

Sarcocystis neurona infections in sea otter (Enhydra lutris): Evidence for natural infections with sarcocysts and transmission of infection to opossums (Didelphis virginiana)

Although Sarcocystis neurona has been identified in an array of terrestrial vertebrates, recent recognition of its capacity to infect marine mammals was unexpected. Here, sarcocysts from 2 naturally infected sea otters (Enhydra lutris) were characterized biologically, ultrastructurally, and genetically. DNA was extracted from frozen muscle of the first of these sea otters and was characterized as S. neurona by polymerase chain reation (PCR) amplification followed by restriction fragment length polymorphism analysis and sequencing. Sarcocysts from sea otter no. 1 were up to 350 I?m long, and the villar protrusions on the sarcocyst wall were up to 1.3 I?m long and up to 0.25 I?m wide. The villar protrusions were tapered towards the villar tip. Ultrastructurally, sarcocysts were similar to S. neurona sarcocysts from the muscles of cats experimentally infected with S. neurona sporocysts. Skeletal muscles from a second sea otter failed to support PCR amplification of markers considered diagnostic for S. neurona but did induce the shedding of sporocysts when fed to a laboratory-raised opossum (Didelphis virginiana). Such sporocysts were subsequently fed to knockout mice for the interferon-gamma gene, resulting in infections with an agent identified as S. neurona on the basis of immunohistochemistry, serum antibodies, and diagnostic sequence detection. Thus, sea otters exposed to S. neurona may support the development of mature sarcocysts that are infectious to competent definitive hosts.

Washington↗

USGS1993

Coupled effects of vertical mixing and benthic grazing on phytoplankton populations in shallow, turbid estuaries

Coastal ocean waters tend to have very different patterns of phytoplankton biomass variability from the open ocean, and the connections between physical variability and phytoplankton bloom dynamics are less well established for these shallow systems. Predictions of biological responses to physical variability in these environments is inherently difficult because the recurrent seasonal patterns of mixing are complicated by aperiodic fluctuations in river discharge and the high-frequency components of tidal variability. We might expect, then, less predictable and more complex bloom dynamics in these shallow coastal systems compared with the open ocean. Given this complex and dynamic physical environment, can we develop a quantitative framework to define the physical regimes necessary for bloom inception, and can we identify the important mechanisms of physical-biological coupling that lead to the initiation and termination of blooms in estuaries and shallow coastal waters? Numerical modeling provides one approach to address these questions. Here we present results of simulation experiments with a refined version of Cloern's (1991) model in which mixing processes are treated more realistically to reflect the dynamic nature of turbulence generation in estuaries. We investigated several simple models for the turbulent mixing coefficient. We found that the addition of diurnal tidal variation to Cloern's model greatly reduces biomass growth indicating that variations of mixing on the time scale of hours are crucial. Furthermore, we found that for conditions representative of South San Francisco Bay, numerical simulations only allowed for bloom development when the water column was stratified and when minimal mixing was prescribed in the upper layer. Stratification, however, itself is not sufficient to ensure that a bloom will develop: minimal wind stirring is a further prerequisite to bloom development in shallow turbid estuaries with abundant populations of benthic suspension feeders.

Journal of Marine Research↗

USGS2005

Sex structure and potential female fecundity in a Epinephelus guttatus spawning aggregation: Applying ultrasonic imaging

Ultrasonic imaging was used to determine the spawning population structure and develop a fecundity estimation model for a red hind Epinephelus guttatus spawning aggregation within the Red Hind Bank Marine Conservation District, St Thomas, U.S.V.I. The spawning population showed considerable within‐month and between‐month variation in population size‐ and sex‐structure. In the spawning season studied, males appeared to arrive at the aggregation site first in December although females represented a large proportion of the catch early in the aggregation periods in January and February. Spawning occurred in January and February, and size frequency distributions suggested that an influx of small females occurred during the second spawning month. An overall sex ratio of 2·9 : 1 (female : male) was recorded for the whole reproductive season. The sex ratio, however, differed between months and days within months. More females per male were recorded in January than in February when the sex ratio was male biased. Fecundity estimates for this species predicted very high potential fecundities (2·4 × 10 5 −2·4 × 10 6 oocytes). The ultrasound model also illustrated a rapid increase in potential female fecundity with total length. Ultrasonic imaging may prove a valuable tool in population assessment for many species and locations in which invasive fishing methods are clearly undesirable.

Journal of Fish Biology↗

USGS2009

Linking marine and freshwater growth in western Alaska Chinook salmon Oncorhynchus tshawytscha

The hypothesis that growth in Pacific salmon Oncorhynchus spp. is dependent on previous growth was tested using annual scale growth measurements of wild Chinook salmon Oncorhynchus tshawytscha returning to the Yukon and Kuskokwim Rivers, Alaska, from 1964 to 2004. First-year marine growth in individual O. tshawytscha was significantly correlated with growth in fresh water. Furthermore, growth during each of 3 or 4 years at sea was related to growth during the previous year. The magnitude of the growth response to the previous year's growth was greater when mean year-class growth during the previous year was relatively low. Length (eye to tail fork, LETF) of adult O. tshawytscha was correlated with cumulative scale growth after the first year at sea. Adult LETF was also weakly correlated with scale growth that occurred during freshwater residence 4 to 5 years earlier, indicating the importance of growth in fresh water. Positive growth response to previous growth in O. tshawytscha was probably related to piscivorous diet and foraging benefits of large body size. Faster growth among O. tshawytscha year classes that initially grew slowly may reflect high mortality in slow growing fish and subsequent compensatory growth in survivors. Oncorhynchus tshawytscha in this study exhibited complex growth patterns showing a positive relationship with previous growth and a possible compensatory response to environmental factors affecting growth of the age class.

Alaska↗

USGS2001

Estimation of population size using open capture-recapture models

One of the most important needs for wildlife managers is an accurate estimate of population size. Yet, for many species, including most marine species and large mammals, accurate and precise estimation of numbers is one of the most difficult of all research challenges. Open-population capture-recapture models have proven useful in many situations to estimate survival probabilities but typically have not been used to estimate population size. We show that open-population models can be used to estimate population size by developing a Horvitz-Thompson-type estimate of population size and an estimator of its variance. Our population size estimate keys on the probability of capture at each trap occasion and therefore is quite general and can be made a function of external covariates measured during the study. Here we define the estimator and investigate its bias, variance, and variance estimator via computer simulation. Computer simulations make extensive use of real data taken from a study of polar bears (Ursus maritimus) in the Beaufort Sea. The population size estimator is shown to be useful because it was negligibly biased in all situations studied. The variance estimator is shown to be useful in all situations, but caution is warranted in cases of extreme capture heterogeneity.

Journal of Agricultural, Biological, and Environme↗

USGS2009

In vitro biology of fibropapilloma-associated turtle herpesvirus and host cells in Hawaiian green turtles (Chelonia mydas)

Fibropapillomatosis (FP) of green turtles has a global distribution and causes debilitating tumours of the skin and internal organs in several species of marine turtles. FP is associated with a presently non-cultivable alphaherpesvirus Chelonid fibropapilloma-associated herpesvirus (CFPHV). Our aims were to employ quantitative PCR targeted to pol DNA of CFPHV to determine (i) if DNA sequesters by tumour size and/or cell type, (ii) whether subculturing of cells is a viable strategy for isolating CFPHV and (iii) whether CFPHV can be induced to a lytic growth cycle in vitro using chemical modulators of replication (CMRs), temperature variation or co-cultivation. Additional objectives included determining whether non-tumour and tumour cells behave differently in vitro and confirming the phenotype of cultured cells using cell-type-specific antigens. CFPHV pol DNA was preferentially concentrated in dermal fibroblasts of skin tumours and the amount of viral DNA per cell was independent of tumour size. Copy number of CFPHV pol DNA per cell rapidly decreased with cell doubling of tumour-derived fibroblasts in culture. Attempts to induce viral replication in known CFPHV-DNA-positive cells using temperature or CMR failed. No significant differences were seen in in vitro morphology or growth characteristics of fibroblasts from tumour cells and paired normal skin, nor from CFPHV pol-DNA-positive intestinal tumour cells. Tumour cells were confirmed as fibroblasts or keratinocytes by positive staining with anti-vimentin and anti-pancytokeratin antibodies, respectively. CFPHV continues to be refractory to in vitro cultivation.

Hawai'i↗

USGS2021

Direct and size-mediated effects of temperature and ration-dependent growth rates on energy reserves in juvenile anadromous alewives (Alosa pseudoharengus)

Growth rate and energy reserves are important determinants of fitness and are governed by endogenous and exogenous factors. Thus, examining the influence of individual and multiple stressors on growth and energy reserves can help estimate population health under current and future conditions. In young anadromous fishes, freshwater habitat quality determines physiological state and fitness of juveniles emigrating to marine habitats. We tested how temperature and food availability affect survival, growth, and energy reserves in juvenile anadromous alewives ( Alosa pseudoharengus ), a forage fish distributed along the eastern North American continent. Field-collected juvenile anadromous A. pseudoharengus were exposed for 21 days to one of two temperatures (21°C and 25°C) and one of two levels of food rations (1% or 2% tank biomass daily) and compared for differences in final size, fat mass-at-length, lean mass-at-length, and energy density. Increased temperature and reduced ration both led to lower growth rates and the effect of reduced ration was greater at higher temperature. Fat mass-at-length decreased with dry mass and energy density increased with total length, suggesting size-based endogenous influences on energy reserves. Lower ration also directly decreased fat mass-at-length, lean mass-at-length and energy density. Given the fitness implications of size and energy reserves, temperature and food availability should be considered important indicators of nursery habitat quality and incorporated in A. pseudoharengus life history models to improve forecasting of population health under climate change.

Journal of Fish Biology↗

USGS2003

Seabird tissue archival and monitoring project: Egg collections and analytical results 1999-2002

In 1998, the U.S. Geological Survey Biological Resources Division (USGS-BRD), the U.S. Fish and Wildlife Service (USFWS) Alaska Maritime National Wildlife Refuge (AMNWR), and the National Institute of Standards and Technology (NIST) began the Seabird Tissue Archival and Monitoring Project (STAMP) to collect and cryogenically bank tissues from seabirds in Alaska for future retrospective analysis of anthropogenic contaminants. The approach of STAMP was similar to that of the Alaska Marine Mammal Tissue Archival Project (AMMTAP). AMMTAP was started in 1987 by NIST and the National Oceanic and Atmospheric Administration (NOAA) as part of the Outer Continental Shelf Environmental Assessment Program sponsored by the Minerals Management Service. Presently sponsored by the USGS-BRD, AMMTAP continues its work as part of a larger national program, the Marine Mammal Health and Stranding Response Program. AMMTAP developed carefully designed sampling and specimen banking protocols. Since 1987, AMMTAP has collected tissues from marine mammals taken in Alaska Native subsistence hunts and has cryogenically banked these tissues at the NIST National Biomonitoring Specimen Bank (NBSB). Through its own analytical work and working in partnership with other researchers both within and outside Alaska, AMMTAP has helped to develop a substantial database on contaminants in Alaska marine mammals. In contrast, data and information is limited on contaminants in Alaska seabirds, which are similar to marine mammals in that they feed near the top of the food chain and have the potential for accumulating anthropogenic contaminants. During its early planning stages, STAMP managers identified the seabird egg as the first tissue of choice for study by the project. There is a relatively long history of using bird eggs for environmental monitoring and for investigating the health status of bird populations. Since 1998, protocols for collecting and processing eggs, and cryogenically banking egg samples have been developed by STAMP (see York et al. 2001). Eggs are being collected on an annual basis for several species at nesting colonies throughout Alaska. Aliquots of these egg samples are being analyzed on a regular basis for persistent organic pollutants and mercury. Results of this work have been published in scientific journals (Christopher et al. 2002) and in conference proceedings (Kucklick et al. 2002; Vander Pol et al. 2002a, 2002b). The intent of this report is to provide an up-to-date description of STAMP. The report contains the most recent egg collection inventory, analytical data, preliminary interpretations based on these data, and a discussion of possible future directions of the project.

Report↗

USGS2020

Regional ocean models indicate changing limits to biological invasions in the Bering Sea

Minimal vessel traffic and cold water temperatures are believed to limit non-indigenous species (NIS) in high-latitude ecosystems. We evaluated whether suitable conditions exist in the Bering Sea for the introduction, survival, and reproduction of NIS. We compiled temperature and salinity thresholds of known NIS and compared these to ocean conditions projected during two study periods: current (2003-2012) and mid-century (2030-2039). We also explored patterns of vessel traffic and connectivity for U.S. Bering Sea ports. We found the southeastern Bering Sea had suitable conditions for the year-round survival of 80% of NIS assessed (n=42). However, only 52% of NIS had conditions suitable for reproduction or development (n=25). Conditions north of 58° N that include sub-zero winter water temperatures were unsuitable for the survival and reproduction of most NIS. While mid-century models predicted a northward expansion of suitable conditions, conditions for reproduction remained marginal. Within the highly suitable southeastern Bering Sea is the port of Dutch Harbor, which received the most vessel arrivals and ballast water discharge in the U.S. Bering Sea. Our findings illustrate the potential vulnerability of a commercially important subarctic ecosystem and highlight the need to consider NIS reproductive and developmental life phases when evaluating limits to their establishment.

ICES Journal of Marine Science↗

USGS2025

Laying sequence and oceanographic factors affect egg size in Scripps's Murrelets Synthliboramphus scrippsi at Santa Barbara Island

Egg size is an important avian life history parameter, with larger eggs indicating greater investment of resources in the chick. Prey availability can affect such investment. We investigated the effects of oceanographic conditions and laying sequence on Scripps's Murrelet Synthliboramphus scrippsi egg size at Santa Barbara Island, California during 2009-2017. We evaluated oceanographic covariates characterizing marine productivity for their effect on egg size, including large-scale oceanographic indices such as the Pacific Decadal Oscillation (PDO) index, Oceanic Niño Index (ONI), and North Pacific Gyre Oscillation (NPGO) index. We also evaluated a larval anchovy catch-per-unit-effort (ANCHL) index and the Biologically Effective Upwelling Transport Index (BEUTI) as region-wide indices, and sea surface temperature (SST) as a local index. We evaluated oceanographic conditions over the entire year and during the breeding season only. We also considered the contribution of lagged effects to oceanographic conditions. Our results generally ran counter to our hypothesis that increased ocean productivity should increase egg size. Based on Akaike's Information Criterion, the four top-ranked models provided support for an association between larger eggs and conditions indicative of lower oceanographic productivity, including lower values of BEUTI and NPGO, and higher values of ONI, PDO, and SST. The only result that supported our hypothesis was a positive relationship between ANCHL and egg size, although the 95% confidence interval for the effect included 0. The strongest relationship detected was between laying sequence and egg size, as second eggs were considerably larger than first eggs. Our results indicate substantial complexity in the relationship between ocean productivity and seabird demography. A better understanding of how ocean productivity affects seabird breeding outcomes through multiple mechanisms will help improve predictions of how seabirds will respond to changing ocean conditions.

California↗

USGS2008

Hierarchical modeling of cluster size in wildlife surveys

Clusters or groups of individuals are the fundamental unit of observation in many wildlife sampling problems, including aerial surveys of waterfowl, marine mammals, and ungulates. Explicit accounting of cluster size in models for estimating abundance is necessary because detection of individuals within clusters is not independent and detectability of clusters is likely to increase with cluster size. This induces a cluster size bias in which the average cluster size in the sample is larger than in the population at large. Thus, failure to account for the relationship between delectability and cluster size will tend to yield a positive bias in estimates of abundance or density. I describe a hierarchical modeling framework for accounting for cluster-size bias in animal sampling. The hierarchical model consists of models for the observation process conditional on the cluster size distribution and the cluster size distribution conditional on the total number of clusters. Optionally, a spatial model can be specified that describes variation in the total number of clusters per sample unit. Parameter estimation, model selection, and criticism may be carried out using conventional likelihood-based methods. An extension of the model is described for the situation where measurable covariates at the level of the sample unit are available. Several candidate models within the proposed class are evaluated for aerial survey data on mallard ducks (Anas platyrhynchos).

Journal of Agricultural, Biological, and Environme↗

USGS1984

Breeding biology of the blue-gray noddy

Blue-gray Noddies, the smallest marine terns, are similar in many respects to all tropical terns in Hawaii: single-egg clutches are laid, growth and development take about 7 weeks, breeding is colonial. Its small size results in eggs that comprise over 27% of adult body weight, compared to 15-20% for most marine terns (Langham 1983). Blue-gray Noddies are widespread in the tropical Pacific, but populations are generally small. This may be the result of its inshore feeding habits and the fact that it is a resident species (Diamond 1978). However, populations in the Hawaiian Archipelago are probably limited by the availability of suitable nest sites in cliffs or rocky outcrops, not food supplies..... Food habits in Hawaii confirm the unique dependence of this species on sea-striders but consumption may be seasonal Blue-gray Noddies take the smallest prey of any seabird in Hawaii and may feed on a lower trophic level..... The Hawaiian population is apparently heavier and produces larger eggs than Blue-gray Noddies elsewhere in the Pacific. This conforms with the general proposition that Hawaiian seabirds are larger than those in the central Pacific (Harrison et al. 1983). The Hawaiian population also has a more predictable breeding season than those farther south.This may be due to a greater seasonality of food supply, but the factors that control the timing of breeding are not clear. There does not appear to be any competition for nest sites with other seabirds.....Our information on growth and development will enable future investigators to estimate the ages of chicks during brief visits to Blue-gray Noddy colonies. This will facilitate programs that are designed to monitor the basic health of seabird populations and to detect changes from baseline that may result from human activities or oceanographic conditions.

Journal of Field Ornithology↗

USGS2015

Ecological effects of the harvest phase of geoduck clam (Panopea generosa Gould, 1850) aquaculture on infaunal communities in southern Puget Sound, Washington USA.

Intertidal aquaculture for geoducks ( Panopea generosa Gould, 1850) is expanding in southern Puget Sound, Washington, where gently sloping sandy beaches are used for field culture. Geoduck aquaculture contributes significantly to the regional economy, but has become controversial because of a range of unresolved questions involving potential biological impacts on marine ecosystems. From 2008 through 2012, the authors used a “before-after-control-impact” experimental design, emphasizing spatial scales comparable with those used by geoduck culturists to evaluate the effects of harvesting market-ready geoducks on associated benthic infaunal communities. Infauna were sampled at three different study locations in southern Puget Sound at monthly intervals before, during, and after harvests of clams, and along extralimital transects extending away from the edges of cultured plots to assess the effects of harvest activities in adjacent uncultured habitat. Using multivariate statistical approaches, strong seasonal and spatial signals in patterns of abundance were found, but there was scant evidence of effects on the community structure associated with geoduck harvest disturbances within cultured plots. Likewise, no indications of significant “spillover” effects of harvest on uncultured habitat adjacent to cultured plots were noted. Complementary univariate approaches revealed little evidence of harvest effects on infaunal biodiversity and indications of modest effects on populations of individual infaunal taxa. Of 10 common taxa analyzed, only three showed evidence of reduced densities, although minor, after harvests whereas the remaining seven taxa indicated either neutral responses to harvest disturbances or increased abundance either during or in the months after harvest events. It is suggested that a relatively active natural disturbance regime, including both small-scale and large-scale events that occur with comparable intensity but more frequently than geoduck harvest events in cultured plots, has facilitated assemblage-level infaunal resistance and resilience to harvest disturbances.

Washington↗

USGS2022

Alaskan Yelloweye Rockfish fecundity revealed through an automated egg count and digital imagery method

Spawning stock biomass (SSB) is often used as an index for reproductive potential (RP) in fisheries stock assessments. This method assumes that mature female biomass is proportional to total egg production and implies that (1) the fecundity–length relationship follows a cubic function or (2) relative fecundity is constant. For many marine fishes, adequate fecundity estimates to evaluate these relationships are lacking. This study estimated fecundity and fecundity relationships for Yelloweye Rockfish Sebastes ruberrimus and evaluated an automated method of counting eggs and larvae. We collected Yelloweye Rockfish ovaries ( N = 90) from the northern Gulf of Alaska, including Prince William Sound, Alaska, during 2018–2019 and used the gravimetric method and image analysis software to count eggs from digital camera images. To evaluate the speed, accuracy, and precision of the automated counting procedure, one-third of the gravimetric samples were also manually counted. Image analysis software was approximately four times faster but equally accurate and precise for fecundity estimates relative to manual counts. Fecundity ranged from 53,249 to 3.052 × 10 6 eggs (mean ± SD = 896,762 ± 699,504 eggs), and relative fecundity increased with female FL and ranged from 68 to 435 eggs/g of body weight (mean ± SD = 226 ± 87 eggs/g). The use of SSB for Yelloweye Rockfish stock assessment could underestimate the contribution to egg production by larger (>5.6-kg) females, overestimate the contribution by smaller females, and lead to biased biological reference points. This study provides critical information to more realistically model RP and improve stock assessment inputs for the development of harvest control rules for Yelloweye Rockfish. Additionally, the use of image analysis software to count eggs in digital images proved to be an effective fecundity estimation method that could be applied to other highly fecund fish species for which the time demand of manual counting methods would be prohibitive.

Alaska↗

USGS2015

Evaluating unsupervised methods to size and classify suspended particles using digital in-line holography

Substantial information can be gained from digital in-line holography of marine particles, eliminating depth-of-field and focusing errors associated with standard lens-based imaging methods. However, for the technique to reach its full potential in oceanographic research, fully unsupervised (automated) methods are required for focusing, segmentation, sizing and classification of particles. These computational challenges are the subject of this paper, in which we draw upon data collected using a variety of holographic systems developed at Plymouth University, UK, from a significant range of particle types, sizes and shapes. A new method for noise reduction in reconstructed planes is found to be successful in aiding particle segmentation and sizing. The performance of an automated routine for deriving particle characteristics (and subsequent size distributions) is evaluated against equivalent size metrics obtained by a trained operative measuring grain axes on screen. The unsupervised method is found to be reliable, despite some errors resulting from over-segmentation of particles. A simple unsupervised particle classification system is developed, and is capable of successfully differentiating sand grains, bubbles and diatoms from within the surf-zone. Avoiding miscounting bubbles and biological particles as sand grains enables more accurate estimates of sand concentrations, and is especially important in deployments of particle monitoring instrumentation in aerated water. Perhaps the greatest potential for further development in the computational aspects of particle holography is in the area of unsupervised particle classification. The simple method proposed here provides a foundation upon which further development could lead to reliable identification of more complex particle populations, such as those containing phytoplankton, zooplankton, flocculated cohesive sediments and oil droplets.

Journal of Atmospheric and Oceanic Technology↗

USGS2023

Isothermal recombinant polymerase amplification and CRIPSR (CAS12A) assay detection of Renibacterium salmoninarum as an example for wildlife pathogen detection in environmental DNA samples

Improving rapid detection methods for pathogens is important for research as we collectively aim to improve the health of ecosystems globally. In the northern hemisphere, the success of salmon ( Oncorhynchus spp.) populations is vitally important to the larger marine, aquatic, and terrestrial ecosystems they inhabit. This has led to managers cultivating salmon in hatcheries and aquaculture to bolster their populations, but young salmon face many challenges, including diseases such as bacterial kidney disease (BKD). Early detection of the BKD causative agent, Renibacterium salmoninarum , is useful for managers to avoid outbreaks in hatcheries and aquaculture stocks to enable rapid treatment with targeted antibiotics. Isothermal amplification and CRIPSR-Cas12a systems may enable sensitive, relatively rapid, detection of target DNA molecules from environmental samples compared to quantitative PCR (qPCR) and culture methods. We used these technologies to develop a sensitive and specific rapid assay to detect R. salmoninarum from water samples using isothermal recombinase polymerase amplification (RPA) and an AsCas12a RNA-guided nuclease detection. The assay was specific to R. salmoninarum (0/10 co-occurring or closely related bacteria detected) and sensitive to 0.0128 pg/µL of DNA (approximately 20–40 copies/µL) within 10 min of Cas activity. This assay successfully detected R. salmoninarum environmental DNA in 14/20 water samples from hatcheries with known quantification for the pathogen via previous qPCR (70% of qPCR-positive samples). The RPA-CRISPR/AsCas12a assay had a limit of detection (LOD) of >10 copies/µL in the hatchery water samples and stochastic detection below 10 copies/µL, similar to but slightly higher than the qPCR assay. This LOD enables 37 C isothermal detection, potentially in the field, of biologically relevant levels of R. salmoninarum in water. Further research is needed to develop easy-to-use, cost-effective, sensitive RPA/CRISPR-AsCas12a assays for rapidly detecting low concentrations of wildlife pathogens in environmental samples.

Journal of Wildlife Diseases↗