Ceratonova shasta Monitoring Studies in the Klamath River

Life cycle of Ceratonova shastaCeratonova 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 polychaete worms infect salmonid fishes and develop into myxospores which then infect polychaetes (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, polychaete 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.

 

 

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
Barth Monitoring fish cages Barth Monitoring trout fry

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).

DATA UPDATES: The first fish exposure for 2019 is scheduled for April 19 - 22 at Beaver Creek (KBC) and Seiad Valley (KSV).

The second fish exposure for 2019 is scheduled for: May 15 - 18 at all index sites.

The third sentinel fish exposure is scheduled for: June 12 - 15 at all index sites.

The final exposure is scheduled for: September 18-21 at Beaver Creek (KBC) and Seiad Valley (KSV).

 

Water Samples

Barth Monitoring filtering
 Barth Monitoring filter paper

 Filtering a water sample
using a vacuum pump

 Folding the filter paper
with captured material

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 ISCO 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.

2019 DATA UPDATES:

Feb 18 - OSU has processed and assayed water samples from Klamath River sites KBC and KSV up to and including 02/18/2019. No C. shasta has been detected yet this year, which is consistent with previous years.

March 25 - no C. shasta DNA was detected in the majority of water samples from KBC and KSV collected through March 25; <1 spore/L was detected in some samples. The first KMN sampling occurred this week and no C. shasta was detected in these.

Expedited processing of water samples will occur April through June. 

April 1 - First week of expedited sample processing. High sediment loads in water samples delayed filtering, shipping and subsequent analyses by one day. Sampling occurred at five index sites: KI5, KBC, KMN (manual sampling), KSV and KOR. Low levels of C. shasta DNA (<1 spore/L) were detected at KI5, KMN and KSV; no C. shasta DNA was detected at the other sites.

April 8 -  Despite higher levels of sediment in water samples, the DNA after processing at OSU did not show signs of inhibiting the PCR under normal protocol conditions. All samples, except KOR, were manual grab samples as ISCO automatic samplers had been pulled in anticipation of higher than normal flows this week. We detected very low levels (<1sp/L) of C. shasta at all sites assayed this week, except KSV at which no C. shasta DNA was detected.

 

 

 Density of Ceratonova shasta in water samples collected at Beaver Creek index site in the Klamath River.
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 2018 (blue data points) compared to 2017 (shaded).

 

 Density of Ceratonova shasta in water samples collected at lower river index sites in 2018

Density (average spores per liter) of Ceratonova shasta in 24-hour composite water samples collected at the mainstem index sites in 2018. 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. 

 

 

Density (average spores per liter) of Ceratonova shasta in 24-hour composite water samples collected at the mainstem index sites Beaver Creek (KBC) and Orleans (KOR) in 2018. 

 

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. 

2019 DATA UPDATES:

Genotyping will commence in April.

 

 Polychaete Sampling

Polychaete sampling

 

Polychaete sampling

Manayunkia 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, polychaete 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.

 

DATA UPDATES:  Polychaete sampling will occur in March at all sites (depending on flow). 

 

 

Polychaete sampling

Model validation sampling

Dr. Alexander collecting a Hess sample at Beaver Creek on SCUBA Field crew 2018 - Ryan Bernstein and Sylvia Gwozdz (USFWS Arcadata office), Robert Predosa (OSU scientific diver volunteer) pictured at Community Center

 

 ANNUAL REPORTS

Bureau of Reclamation Report for 2008

Bureau of Reclamation Report for 2009 

Bureau of Reclamation Report for 2010

Bureau of Reclamation Report for 2011

Bureau of Reclamation Report for 2012

Bureau of Reclamation Report for 2013

Bureau of Reclamation Report for 2014

Bureau of Reclamation Report for 2015

Bureau of Reclamation Report for 2016

Bureau of Reclamation Report for 2017

B

 '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.