OREGON STATE UNIVERSITY
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Ceratonova shasta is a freshwater, myxozoan parasite that is native to the Pacific North West of North America. It causes enteronecrosis in juvenile salmonids and is associated with population-level impacts in the Klamath River. Transmission occurs through waterborne stages: actinospores released from annelids infect salmonid fishes and develop into myxospores which then infect annelids (see life cycle on right). The parasite proliferates in each host.
In response to the high prevalence and severity of C. shasta-infection in Klamath salmonids, we developed a parasite monitoring program to track the spatial and temporal abundance of C. shasta. The three main approaches are based on the parasite's life cycle and include sentinel fish exposures, annelid host sampling and molecular quantification of parasite DNA in water samples. These are described in more detail below. Monitoring occurs at established index sites which are shown on the following map.
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Klamath River Index Sites with site abbreviations and river kilometers (Rkm). Iron Gate Dam (Rkm 306) blocks anadromous salmonid migration. |
Sentinel Fish Exposures
| Sentinel fish cages in the Klamath River |
Susceptible out- of-basin rainbow trout |
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In sentinel fish exposures, fish highly susceptible to the parasite (out-of-basin rainbow trout) are placed in cages alongside fish of interest such as in-basin Chinook and coho salmon at index sites along the river for a three day exposure. All fishes are transported to OSU's John L Fryer Aquatic Animal Health Lab and monitored for infection (~ 60 days). Severity of infection (percent morbidity and mean days to morbidity) are recorded through visual observations and molecular assay (PCR).
2021 DATA UPDATES: Field work reamins subject to COVID-restrictions.
An April sentinel fish exposure occurred at KBC and KSV 04/19-04/22 with IGH Fall Chinook and out of basin rainbow trout. Updates can be accessed here.
The May sentinel fish exposure occurred 17th-20th. Report
In addition, an experiment to compare the relative susceptibility of two Chinook salmon stocks, IGH Fall Chinook and TRH Spring Chinook, occurred May 24-27th. Report
The June sentinel fish exposure occurred June 14th-17th.
The September (and final for 2021) sentinel fish exposure occurred September 20th - 23rd. Chinook and out of basin rainbow trout were held at KBC and KSV.
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| Developmental stages of Ceratonova shasta in an intestinal smear from a fall Chinook exposed at KSV in April 2019. Note the pseudopodia extending from the pansporocyst and the cells within cells. Two myxospores will develop within this 'disporoblast' stage. | Developmental stages of Ceratonova shasta, including an immature myxospore, in a kidney smear from a fall Chinook exposed at KSV in April 2019. The infection was fatal. |
Water Samples
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Filtering a water sample |
Folding the filter paper |
To detect and quantify waterborne stages of C. shasta, river water samples are collected at five mainstem index sites; once a week all year round at two sites (KBC and KSV) and once per week from April through October at three other sites. Solar-powered automatic samplers (ISCOs) collect 1L water every 2 hours for 24 hours, from which 4 1L samples are manually taken. Each 1L sample is filtered through a nitrocellulose membrane using a vacuum pump, and any captured DNA in 3 of the replicate samples is extracted using a kit. A quantitative PCR (qPCR) specific for C. shasta is used to detect and quantify any parasite DNA present. Cq values generated by the qPCR are converted to numbers of parasite spores per liter of water using reference samples with known quantities of spores. The Karuk and Yurok tribal biologists are integral to the collection and filtration of the water samples. Water samples are also taken in conjunction with the sentinel fish exposures; manual 'grab' samples are collected on the first and last day of the exposure. Data are presented as the average spores per liter of three replicate 1 liter water samples collected at each site and time.
2021 DATA UPDATES:
During Winter, sampling occurs only at two index sites, KBC and KSV. Spatial sampling will increase to six sites in Spring.
1/19/2021. Both KBC and KSV were negative for C. shasta in the January samples.
KOR 2 spores/L
3/29/2021: C. shasta DNA was detected at 5 out of 6 sites. Density has increased:
KOR 2 spores/L
KTC 1 spore/L
4/05/2021: Expedited sample processing has commenced. C. shasta DNA was detected at all 6 index sites. Density increased at all sites, an order of magnitude at two sites, KMN and KSV. All samples were collected with an ISCO.
KOR 7 spores/L
KTC 4 spores/L
KOR 18 spores/L
KTC 14 spores/L
04/19/2021: The change in density of waterborne C. shasta varied by site. The highest density was again at KBC with 79 spores per liter measured. Density doubled at one site, KI5 but decreased at the remaining 4 sites (halved at 2 sites).
KOR 12 spores/L
KTC 6 spores/L
04/26/2021: The density of waterborne C. shasta increased (doubled) only at the uppermost index site, KI5. Density remained the same at the two lowermost sites, KOR and KTC and decreased at the remaining three sites in between.
KI5 61 spores/L
KBC 34 spores/L
KMN 32 spores/L
KSV 4 spores/L
KOR 12 spores/L
KTC 5 spores/L
05/03/2021: The densities of waterborne C. shasta were similar this week to those last week. A clear infectious zone continued to span the uppermost index sites (KI5 to KMN). The highest density measured was an average of 55 spores per liter at KI5. All samples were collected with an ISCO May 2-3, with the exception of KOR which were manually collected on May 3.
KI5 55 spores/L
KBC 36 spores/L
KMN 29 spores/L
KSV 3 spores/L
KORg 10 spores/L
KTC 7 spores/L
05/10/2021: We measured a decrease in density of 3-6 fold at the three uppermost sites: KI5, KBC & KMN, changing from 29-55 spores per liter down to 6-9 spores per liter. Densities at the lowermost sites (2-10 spores per liter) were similar to last week. There remains a clear decrease in abundance from KMN to KSV. Density decreased at all sites except KTC which increased by 43%. All samples were collected with an ISCO May 9-10.
KI5 8 spores/L
KBC 6 spores/L
KMN 9 spores/L
KSV 2 spores/L
KOR 7 spores/L
KTC 10 spores/L
05/17/2021: Spore densities increased at 5 out of 6 sites, to > 10spores per liter at 4 of those, including KTC. We measured a 3-7 fold increase in spore density at the three uppermost sites: KI5, KBC & KMN, changing from 6-9 spores per liter up to 21-47 spores per liter.
KI5 33 spores/L
KBC 47 spores/L
KMN 21 spores/L
KSV 4 spores/L
KOR 6 spores/L
KTC 13 spores/L
05/24/2021: Spore densities of waterborne C. shasta increased at some sites and decreased at others. Spore densities exceeded 10 spores per liter at 4 of the 5 sites. The highest density measured was at KI5 with 36 spores per liter. Notably, spore densities at KOR were three times higher this week, with 21 spores per liter measured, the highest detected there this season.
KI5 36 spores/L
KBC 34 spores/L
KMN 13 spores/L
KSV 6 spores/L
KOR 21 spores/L
KTC 5 spores/L (grab)
05/31/2021: Spore densities of waterborne C. shasta were lower at 5 of the 6 index sites this week, being about half or more than half of the density measured the previous week at 3 of those sites. The exception was at KSV where density trippled from last week (from 6 to 18 spores per liter).
Total density exceeded 10 spores per liter at 4 sites, however, at those sites 37 – 72.2% of the sample was comprised of genotype II. Thus, the Chinook 40% mortality threshold of 10 spores of genotype I per liter was likely only met at KBC. In contrast, the coho 40% mortality threshold of 5 type II spores per liter was met at 4 index sites, where type II spore density ranged from 5 to 13 spores per liter. Spore density was relatively high for genotype II plus it comprised a relatively high proportion of total spore density.
Site, total spores, (type II), proportion II/total
KI5 18 (12) grab 66.7%
KBC 16 (6) 37.5%
KMN 11 (5) 45.5%
KSV 18 (13) 72.2%
KOR 7 (<1) 14.3%
KTC 4 (0) 0%
06/07/2021: OSU received and assayed this week's water samples from all index sites. All samples except KI5 and KTC were taken by ISCO automatic sampler and are given as the average density of C. shasta spores per liter of river water (from 3x 1-L subsamples of a 24-hour composite sample, Sunday-Monday June 6-7); KI5 and were 3x 1-L manual grab samples taken Monday June 7. Total spore densities fluctuated among sites between the last two weeks. More than 10 spores per liter were measured at 3 sites (uppermost), and after accounting for the contribution of type II, these are also likely above 10 spores per liter of type I (Chinook-relevant). The highest density, of 61 spores per liter, was measured at KI5; note that this was a grab sample rather than a 24-hour composite. Of note: KI5 - total density was 3 times higher this week, and type II was twice as high (see genotyping section below). KSV – density much lower this week - C. shasta was barely detected there; (versus 18 spores per liter the week before)
KI5 8
KBC 4
KMN concluded for the year
KSV not detected
KOR <1
KTC 5
KI5 28
KBC 4
KSV 11
KOR 0
KTC <1
KI5 20
KBC 4
KSV 3 g
KOR 0
KTC <1 g
Site, total spores per liter
KI5 5
KBC not detected
KSV not detected
KOR not detected
KTC <1 ISCO & grab
07/26/2021: A higher density of C. shasta was measured at several sites this week; the most being 5 spores per liter at KI5.
Site, total spores per liter
KI5 5
KBC 3
KSV 0
KOR <1
KTC 2 ISCO, 6 GRAB
08/02/2021: An increase in spore density was measured at KBC and KSV this week, with over 10 spores per liter measured at KBC.
Site, total spores per liter
KI5 2
KBC 14
KSV 8 GRAB
KOR <1
KTC <1
08/09/2021
Site, total spores per liter
KI5 61
KBC 23
KSV 14
KOR 7
KTC 4
08/16/2021
Site, total spores per liter
KI5 36
KBC 4
KSV <1
KOR 9
KTC 26
08/23/2021
Site, total spores per liter
KI5 not collected (wildfires)
KBC not collected
KSV not collected
KOR 9
KTC 13
08/30/2021
Site, total spores per liter
KI5 60
KBC 38
KSV 16
KOR 10
KTC 20
09/07/2021
Site, total spores per liter
KI5 25
KBC 14
KSV 10
KOR 9
KTC not yet determined
09/13/2021
KI5 30 grab
KBC 14 ISCO
KSV 15 grab
KOR 8 ISCO
KTC 12 ISCO
09/20/2021: Densities of C. shasta were similar or higher two weeks ago (9/13) than the preceding week and were remarkably higher again this week (9/20); >10 spores per liter were measured at each site. These levels are higher than usual for this time of year. A mix of collection methods was used but the same method was used at a given site both weeks; the water samples are usually collected at the same time each week at a site and thus should be comparable. Grab samples are often about twice the density of a 24-hour ISCO sample.
KI5 50 grab
KBC 31 ISCO
KSV 20 grab
KOR 13 ISCO
KTC not yet available
10/11/2021: Densties of C. shasta were lower at most sites this week compared to levels three weeks ago (09/20). However, they were still above the usual levels for this time of year with 10 spores per liter measured at 3 out of 5 index sites; densities were highest at KTC. Samples were collected manually ('grab') at KI5 (and both ISCO and grab at KTC).
KI5 10 grab
KBC 10 ISCO
KSV 3 ISCO
KOR 15 ISCO
KTC 40 ISCO, 27 grab
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| Density (average spores per liter) of Ceratonova shasta in 24-hour composite water samples collected at the mainstem longterm index site, near Beaver Creek (KBC), in 2021 (blue data points) compared to 2020 (purple), 2019 (light blue hashed shading), 2018 (yellow dotted shading) and 2015 (gray shading). |
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Density (average spores per liter) of Ceratonova shasta in 24-hour composite water samples collected at the mainstem index sites in 2021. Note that KMN is sampled only during salmonid outmigration, KBC and KSV year round and remaining sites April through October. KBC = near Beaver Creek, KSV = Seiad Valley, KI5 = near I5 bridge, KTC = Tully Creek, KMN = Kinsman Fish Trap, KOR = Orleans. The line denotes 10 spores per liter which corresponds with 40% mortality threhold in Chinook salmon. |
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Density (average spores per liter) of Ceratonova shasta in 24-hour composite water samples collected at the mainstem index sites in 2020. Note that KMN is sampled only during salmonid outmigration, KBC and KSV year round and remaining sites April through October. KBC = near Beaver Creek, KSV = Seiad Valley, KI5 = near I5 bridge, KTC = Tully Creek, KMN = Kinsman Fish Trap, KOR = Orleans. |
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Klamath River Index site KI5 Photos courtesy of Karuk Tribal Biologist Larry Alameda |
KMN - Kinsman Trap | KSV - Seiad Valley | KOR - Orleans |
Genotyping: There are multiple genetic types or genotypes of C. shasta simultaneously present in the Klamath River. These differentially disease the various salmonid species. For example, type I causes mortality in Chinook salmon whereas type II can be fatal for coho salmon. Type 0 is found in sympatric Oncorhynchus mykiss (steelhead and redband rainbow trout).
Therefore, we also genotype each water sample using a qPCR that amplifies the variable ITS1 region and then we sequence that amplicon. From the sequencing chromatogram, we can determine the proportion of each genotype present in a sample. We use the total spore density to then determine the number of spores of each genotype in a sample.
2021 DATA UPDATES:
Genotype data will become available once spore densitities reach 1 spore per liter.
04/05/2021: No coho type II was detected at any index site.
04/12/2021: The coho-relevant gentoype II was detected only at KTC, at a level of 1 spore per liter.
04/19/2021: The coho-relevant gentoype II was detected at three index sites, with the highest density of 3 spores per liter measured at KMN.
Site, total spores, type II
KI5 32, 0
KBC 79, <1
KMN 51, 3
KSV 17, 0
KOR 12, 0
KTC 6, 2
04/26/2021: The coho-relevant genotype II was detected in low densities at five index sites, with the highest density of 2 spores per liter measured at KOR.
Site, total spores, type II
KI5 61 (<1)
KBC 34 (<1)
KMN 32 (1)
KSV 4 (0)
KOR 12 (2)
KTC 5 (<1)
05/03/2021: The coho-relevant genotype II was detected at 6 index sites this week. The highest density measured was 4 spores per liter at KBC and KMN.
Site, total spores, type II
KI5 55 (2)
KBC 36 (4)
KMN 29 (4)
KSV 3 (<1)
KOR g 10 (1)
KTC 7 (1)
05/10/2021: The coho-relevant genotype II was detected at 6 index sites. Densities were low (1 spore per liter).
Site, total spores, type II
KI5 8 (1)
KBC 6 (1)
KMN 9 (1)
KSV 2 (<1)
KOR 7 (1)
KTC 10 (1)
05/17/2021: The coho-relevant genotype II was detected at all 6 index sites. Densities increased at 5 of the 6 sites measured. They were still low at the three lowermost sites (2 spores per liter), but were notably higher at the uppermost three sites being > 5 spores per liter at two of those (6-8 spores per liter at KI5 and KBC). In sentinel coho exposures, > 5 spores per liter corresponds to a 40% mortality threshold.
Site, total spores, (type II)
KI5 33 (6) 18%
KBC 47 (8) 17%
KMN 24 (4) 19%
KSV 4 (<1)
KOR 6 (2) 33%
KTC 13 (2) 15%
05/24/2021: The coho-relevant genotype II was detected at all 5 index sites analysed. The density of genotype II in the upper river samples (KI5 & KBC) remains high this week (at or above 5 spores per liter).
Site, total spores, (type II)
KI5 36 (7)
KBC 34 (5)
KMN 13 (1)
KSV 6 (1)
KOR 21 (1)
KTC 5 (1) grab
05/31/2021: The coho-relevant genotype II was detected at 5 of the 6 index sites. The coho 40% mortality threshold of 5 type II spores per liter was met at 4 index sites, where type II spore density ranged from 5 to 13 spores per liter. Spore density was relatively high for genotype II plus it comprised a relatively high proportion of total spore density.
Site, total spores, (type II), proportion II/total
KI5 18 (12) grab 66.7%
KBC 16 (6) 37.5%
KMN 11 (5) 45.5%
KSV 18 (13) 72.2%
KOR 7 (<1) 14.3%
KTC 4 (0) 0%
06/07/2021: Genotype II was again detected throughout the system and levels exceeded the 5 spore per liter threshold for coho (40% mortality) at 3 sites (uppermost), with up to an average of 22 spores per liter measured at KI5 (grab sample, not ISCO). Because type II densities are crossing the 40% coho mortality threshold, we have included a type II graph also this week. Genotype II (relevant for coho) numbers given in parentheses.
Site, total spores, (type II), proportion II/total
gKI5 61 (22) 37%
KBC 20 (8) 40%
KMN 16 (6) 37%
KSV <1 (<1)
KOR 8 (2) 25%
gKTC 8 (<1)
06/14/2021: Genotype II was most prevalent at the uppermost three sites, with over 5 spores per liter detected at both KI5 and KBC. Almost half the C. shasta at KI5 was comprised of type II.
Site, total spores, (type II), proportion II/total
KI5 37 (17) 46%
KBC 37 (9)
KMN 16 (4)
KSV 0 (0)
KOR 6 (<1)
KTC 14 (<1)
06/21/2021: Genotype II was most prevalent at the uppermost two sites, with over 5 spores per liter detected at both KI5 and KBC. More than half the C. shasta at these two sites was comprised of type II.
Site, total spores, (type II), proportion II/total
KI5 43 (32) 74%
KBC 14 (9) 64%
KMN concluded for the year
KSV <1 (0)
KOR 7 (<1)
KTC delayed
06/28/2021: The density of genotype II was lower this week; below 5 spores per liter at all sites measured. It was most prevalent at the uppermost two sites, with 4 and 2 spores per liter detected at KI5 and KBC, respectively. This constituted half the total C. shasta measured at those sites.
Site, total spores, (type II)
KI5 8 (4)
KBC 4 (2)
KMN [2021 sampling concluded]
KSV not detected (not detected)
KOR <1 (not detected)
KTC 5 (not yet assayed)
07/06/2021: The density of genotype II fluctauted among index sites. 5 spores per liter were detected at KI5, and fewer at other sites.
Site, total spores, (type II)
KI5 28 (5)
KBC 4 (1)
KSV 11 (3)
KOR 0 (0)
KTC <1 (0)
07/12/2021: Samples at KI5, KBC & KOR were taken by an ISCO automatic sampler and are given as the average density of C. shasta spores per liter of river water (from 3x 1-L subsamples of a 24-hour composite sample, Sunday-Monday July 11-12); KSV and KTC were 3x 1-L manual grab samples taken Monday July 12. Genotype II (relevant for coho) numbers given in parentheses
Site, total spores, (type II)
KI5 20 (4)
KBC 4 (<1)
KSV 3 (<1) grab
KOR 0 (0)
KTC <1 (0) grab
07/19/2021: Genotype II was only detected at one site, KI5 with 2 spores per liter measured. This comprised 40% of the total C. shasta measured.
Site, total spores per liter
KI5 5 (2)
KBC not detected (n.d.)
KSV not detected (n.d.)
KOR not detected (n.d.)
KTC <1 (0) grab & ISCO
07/26/2021: Genotype II was detected at three sites, with the highest density of 2 spores per liter measured at KI5.
Site, total spores per liter
KI5 5 (2)
KBC 3 (1)
KSV 0 (0)
KOR <1 (0)
KTC 2 (<1) ISCO, 6 (<1) GRAB
07/02/2021: Genotype II was detected at the three sites tested, with the highest density of 3 spores per liter measured at KBC.
Site, total spores per liter
KI5 2 (...)
KBC 14 (3)
KSV 8 (2) GRAB
KOR <1 (<1)
KTC <1 (...)
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| Density (average spores per liter) of Ceratonova shasta genotype II in 24-hour composite water samples collected at the mainstem index sites in 2021. Note that KMN is sampled only during salmonid outmigration, KBC and KSV year round and remaining sites April through October. KBC = near Beaver Creek, KSV = Seiad Valley, KI5 = near I5 bridge, KTC = Tully Creek, KMN = Kinsman Fish Trap, KOR = Orleans. The line denotes 5 spores per liter which corresponds with a 40% mortality threhold in coho salmon. |
Annelid Sampling
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| Manayunkia occidentalis tubes from the image on right | A high density assemblage sample at TOH |
To monitor abundance and prevalence of infection in the invertebrate host of C. shasta, Manayunkia occidentalis, annelid samples are collected annually at seven sites in the Klamath River spanning a discharge gradient. Sites are located in the upper basin downstream from Keno dam, 3 sites are located in the middle basin downstream from Iron Gate Dam, and 2 sites are located in the lower basin downstream from the Scott and Salmon Rivers. Samples are collected once each in fall, winter, spring, and summer, and more frequently if flooding or pulse flow events are scheduled to occur. Samples are processed to determine density, simple demographics, and prevalence of C. shasta infection.
2021 DATA UPDATES:
Annelid sampling commenced mid-March. Additional sampling will occur the last two weeks of May.
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Annelid sampling can be challenging during high flows. |
HSU diver Dr. Eve Robinson prepares to take substrate data during annelid model project, July 2019 |
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| OSU diver, Dr. Julie Alexander, prepares to collect annelids from the benthos during polychaete model project, July 2019 |
Water quality was poor in July 2019; algae limited visibility |
ANNUAL REPORTS
Annual reports for Bureau of Reclamation funded studies for 2015 onwards are available. Please contact Sascha Hallett (hallettsAToregonstate.edu). Annual reports are submitted June 1 the year after the research year.
ANNUAL KLAMATH RIVER FISH HEALTH WORKSHOP
The latest workshop was held virtually via Zoom on Tuesday March 16th, 2021.
Graphical abstracts from the workshop.
Redescription of the invertebrate host:
For a lyrical rendition with more information, enjoy: The Ballad of Ceratonova shasta
'Tully Creek' Longitudinal Water Sampling
Parasite levels at our lowermost index site, Tully Creek (at ~60 Rkm), have increased over the past several years to surpass those detected at previously high sites, such as Beaver Creek (KBC). To investigate the extent of the high parasite densities at the Tully Creek site, we sampled water downstrea m (beginning at the lowermost road-accessible location, ~38.4Rkm) and upstream of the index site at Orleans (~90Rkm). Link to report.
Data shared here are preliminary and subject to modification.
Page photo credits: S Atkinson, S Hallett & J Alexander
Monitoring Studies are Primarily Supported by the Bureau of Reclamation.
