June 2022 - 17

to make one sample pool for testing. After
samples are collected, they are transported
back to the lab for processing.
During processing the kidney/spleen
samples are homogenized together using a
small rolling pin. This same process is used
for the viscera and whole fish bodies. The
samples are then diluted with a medium,
placed in a machine referred to as a stomacher
to thoroughly combine them and then poured
into sterile tubes. At this point samples are
ready for testing. The kidney/spleen sample is
used to screen for viruses and bacteria, and the
ovarian fluid sample is used for virus testing.
During virus testing a sub-sample is
placed onto a live layer of fish cells within
a plastic flask. These inoculated cells are
incubated and monitored for 28 days for any
virus-like signs that the cells may exhibit.
Viruses commonly screened for include
viral hemorrhagic septicemia virus (VHSV),
infectious pancreatic necrosis virus (IPNV)
and infectious hematopoietic necrosis virus
(IHNV).
Lab personnel commonly screen for three
bacteria pathogens. Two of them include
Yersinia ruckeri, which causes enteric red
mouth disease and Aeromonas salmonicida,
which causes furunculosis. Samples are
streaked onto bacteriology agar in petri dishes,
which are then incubated and monitored for
growth. Bacteria colonies that grow are put
through a series of tests to rule out any suspect
pathogens.
The third bacterial pathogen the lab tests
for is difficult to grow. This bacterium is
Renibacterium salmoninarum and causes
bacterial kidney disease. To test for this, kidney
tissue is smeared onto a microscope slide and
then stained with a fluorescent antibody stain.
This causes the bacteria to glow green when
viewed with a special microscope equipped
with an ultraviolet light bulb.
The heads that are collected during a fish
health inspection are tested for the parasite
Myxobolus cerebralis, which causes whirling
disease. Whirling disease spores reside in the
cartilage and bones of the head. In order to
look for these spores, the fish heads are placed
in a hot water bath overnight. They are then
filtered through a mesh screen to separate the
cartilage and bone from the tissue. Then the
samples are put through various processes
to break apart the bone and cartilage, which
releases and isolates the spores. Samples are
examined under a microscope to screen for
the spores.
The head from a rainbow trout is placed into a tube to test for Myxobolus cerebralis, a parasite that causes whirling
disease. The fish came from Clark's Fork Fish Hatchery as part of an annual fish health certification inspection. The
spores of Myxobolus cerebralis are large enough to view under a microscope after being isolated from the cartilage of
the fish head. (WGFD photo)
Fortunately, pathogens are rarely found
in any of the state's hatcheries. " The fish in
Wyoming's hatcheries are relatively healthy, "
Smith said. " Wyoming is a headwater state
so we get access to the water first. Most of
the hatcheries use either spring water or well
water, or a combination of both, so there is
less possibility of contamination. "
The lab also works on diagnostic cases.
These are cases where fish are sick and dying
at a fish hatchery or in the wild. " We see a
lot of bacterial infections, specifically bacterial
coldwater disease, " Smith said.
Bacterial coldwater disease occurs naturally
in the water. If fish hatchery staff notice sick
fish, they collect fish to send to the lab for
testing. Bacterial pathogens and external
parasites are normally the cause for these
events, and a variety of treatments can be
subscribed, including specially medicated food.
Lab staff often diagnose the cause of
disease fairly quickly, based on symptoms,
bacteria and parasites observed and bacteria
grown in petri dishes so hatcheries don't have
to wait too long to start treatment.
- Robin Kepple, Game and Fish information and education
specialist in the Laramie Region, and Carl Smith,
forensic/fish health biologist, contributed to this story.
Wyoming Wildlife | 17

June 2022

Table of Contents for the Digital Edition of June 2022