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).

 

 

 

 

 

 

 Sentinel Fish Exposures

Barth Monitoring fish cagesSentinel fish cages
in the Klamath River
Barth Monitoring trout frySusceptible
out- of-basin
rainbow trout

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 the lab (AAHL) and monitored for infection (~ 60 days). Prevalence of infection and severity of infection (percent morbidity and mean days to morbidity) are recorded through visual observations and molecular assay (PCR).

The first fish exposure in 2017 occured: April 20 - 23 at Beaver Creek (KBC) and Seiad Valley (KSV).

 

DATA UPDATES: summaries of sentinel fish exposures are as follows:  April  (no fish with disease signs)  May     June   September.

 

 

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 samples are collected on the first and last day of the exposure.

2017 DATA UPDATES: Expedited processing of water samples commenced the first week of April. If ISCO samples are unavailable at time of collection, manual 'grab' samples at one time point (versus 24 h) are collected instead; these are denoted with a 'g'.

April 03: No C. shasta was detected in water samples collected from KBC, KSV and KOR.

April 10: KBC - trace detection of C. shasta in one of the three 1 L replicates; KSV - <1 sp/L detected in all three 1 L replicates; KOR - detection of C. shasta at <1sp/L in one 1L replicate only.

April 17: KBC - no C. shasta was detected in any of the three 1 L grab sample replicates; KSV - trace detection of C. shasta in one of the three grab sample 1 L replicates.

April 24: KBC - trace levels of C. shasta were detected; KSV - >1 but < 5 spores/L of C. shasta detected; KOR - ~5 spores/L of C. shasta detected.

May 1: KBC <1 spore/L; gKSV 1 spore/L; KOR <1 spore/L.

May 8: KBC <1 spore/L; KMN <5 spores/L; KSV <1 spore/L; gKOR <1 spore/L.

May 15: gKBC <1 spore/L; KMN 1 spore/L; KSV 3 spores/L; KOR 2 spores/L.

 

 

 

Density of Ceratonova shasta in Klamath River water samples collected at KBC 2017
Density (spores per liter) of Ceratonova shasta in water samples collected at the mainstem longterm index site, near Beaver Creek (KBC), in 2017 (blue data points) compared to 2016 (shaded).

 

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 O. 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. 

2017 DATA UPDATES:

April 3: No C. shasta was detected at KBC, KSV, or KOR, therefore no genotyping was performed.

April 10: Less than 1 spore/L of C. shasta was detected at KBC, KSV, and KOR, therefore no genotyping was performed.

April 17: Less than 1 spore/L of C. shasta was detected at KBC and KSV, therefore no genotyping was performed.

April 24: Less than 1 spore/L of C. shasta was detected at KBC but >1 spore at KSV and KOR, therefore only KSV and KOR samples were genotyped. Only type I was detected in these samples (if type II was present it would be detected).

May 1: All samples (5) with 1 spore/L detected were genotyped. Type I was present at all three sites - KBC, KSV and KOR. Type II was detected at KSV.

May 8: only genotype I was detected in each of the KMN samples.

May 15: underway.

 

 

 Polychaete Sampling

BARTH polychaete monitoring 

BARTH polychaete monitoring 2

Close up photo showing polychaetes in tubes comprised of mucus and fine sediment grains. Polychaetes are suspension feeders that consume small particles transported by river currents. Ceratonova shasta myxpospores are likely consumed by filter feeding polychaetes. Divers observing polychaetes on boulder substrate in the Klamath River.

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 samples will be collected once river flows decrease to safe levels.

 

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

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

Monitoring Studies are Primarily Supported by the Bureau of Reclamation.