The role of nutritional research in aquaculture Ronald Hardy

The role of nutritional research in aquaculture Ronald Hardy

November 8, 2019 0 By William Morgan


STEPHEN WATTS: Our next speaker today is going
to be Ron Hardy.
Ron is an expert in production fish aquaculture
and has worked in the nutrition of production
fish for years.
He has an incredible expertise in this area.
He came from the University of Idaho, and
you’re still pretty active there these days,
except for the folks here at the San Juan
Islands, I understand.
RONALD HARDY: Which I should be there right
now.
STEPHEN WATTS: You should be there right now.
One of the things that we wanted to bring
into this session was the fact that production
fisheries or production aquaculture has been
studying nutrition a long time.
It’s paramount to the success of those industries
that we understand … the nutritional needs
of fish that you’re going to try to eventually
put on your plate and eat.
That being the case, that expertise can be
transferred even into small fish, which we
generally won’t eat a zebrafish, but a lot
of that information can be transferred, and
Ron agreed to come and tell us a little bit
about the history of some of the production
fish and perhaps give us some insight into
moving forward.
Ron?
Thank you.
STEPHANIE MURPHY: Just a reminder, these little
black pop-up things are your microphones.
So, you might have to, like, cover them up.
RONALD HARDY: Thank you, Steve.
The topic that I was given and Steve described—and
thank you for inviting me, by the way—was
the role of nutrition research in developing
production aquaculture and how this might
be applied to zebrafish, as he said.
Alright, Ron, let’s see here.
I’m off to a good start here, aren’t I?
Alright.
STEPHANIE MURPHY: The other way?
RONALD HARDY: Okay.
Now I’m in business.
So, I thought I’d begin by just giving you
a slide here showing you the growth of aquaculture
over the last few decades.
The reason I’m showing you this is to illustrate
how rapidly it’s grown, but also when you
look at these numbers, and I’ll explain
in a second, realize that this growth has
been supported by development of diets and
it greatly increases …
STEPHANIE HARDY: Excuse me … microphone.
RONALD HARDY: Oh, sorry about that.
I can’t see my slides very well from here.
It’s accompanied by a tremendous increase
in the amount of feed produced by the aquaculture
industry.
So, just to explain this slide a little bit.
What you see in the increasing numbers there
is the amount of fish that are captured in
the wild, which is illustrated by the white
and the green areas, and then the growth of
aquaculture over the last 20 or 30 years in
the red.
You have to discount the part at the bottom—the
green area there.
It refers to industrial fishing, in other
words, fish that are caught and not used directly
for human consumption; and then the purple
line shows the per capita fish supplied worldwide.
So, you see there’s been an upward trend,
and you can also see that over the last … oh,
since about 1990, aquaculture’s proportion
of the amount of seafood or fisheries products
that are consumed or produced for human consumption
has really grown wild, whereas the amount
that’s harvested from the sea has been pretty
flat.
So, we’ve pretty much tapped out all of
the resources that we harvest from the ocean.
Another way of showing the same data that’s
a little bit different slant here.
Aquaculture, we like to go around and tell
everybody—it doesn’t really a result of
any big monies being sent our way, but anyway—it’s
been the most rapid-growing livestock production
sector in the world for the last 20 or 30
years.
Since 2000, FAO says it’s grown 5.8 percent
per year; it grew a little faster before that.
And, as I said, in the yellow there you can
see capture fishery’s pretty much been level
since the 1990s, and aquaculture has increased
tremendously—just taken off like gangbusters.
The latest data shows about 76.6 million metric
tons of fish.
Now, aquaculture includes seafood, shellfish,
and other things, but that’s just fish,
so that’s really gone up crazy.
Why has this happened?
Well, there are a number of reasons.
I’d like to say it’s all nutrition, but
it isn’t.
Vaccines have been a great development, in
that they’ve eliminated or at least reduced
the losses of fish to environmental and bacterial
diseases.
We’ve seen a lot of genetic improvement
in some species, particularly salmon and tilapia,
selecting for growth and other performance.
Government policies—this really refers here
to China.
If you look at the little circle on the bottom,
you can see China accounts for almost two-thirds
of all of the fish production in the world,
and if you see North America there, we’re
at 1.2 percent; we’re not very big.
But—and this is my big “but” here—better
feeds and nutritional requirements.
That’s played a huge role in the development
of aquaculture and expansion.
So anyway, early on—I’m going to give
you a little history here—fish nutrition
research was really based upon not a lot of
fish but mainly the need to produce salmon
and trout for what we call “enhancement
hatcheries.”
So, these are hatcheries that grow little
baby fish, release them into the wild, and
off they go in the ocean to complete the life
cycle, come back, enter the fisheries, and
are captured and contribute to economic activity.
Only in the last 30 years, when farming really
took off, has the need for really well-formulated
feeds taken place.
And … if you think about it, step back … fish
food production, this growth of aquaculture,
has really changed the type of research that
we do.
When I started in this business all we cared
about was producing little baby fish—smolts
and trout—for enhancement and release.
And now, with food production, not only has
the thrust changed, but also the economics
of production has changed, too—the reasons
and the goals of production.
The goals of production used to be, “Let’s
produce a really healthy fish that’s going
to survive after it’s released, we don’t
care what it costs, we only do this once a
year.”
The big goal was just really not lose any
fish and make sure they’re healthy.
Now: let’s make money.
That’s really what the name of the game
is.
So, a little historical background.
We’re all researchers.
We all chase grants.
In 1924, these two gentleman, Embody and Gordon,
did, I think, one of the most fun research
projects—at least it sounds like fun—that
has ever done.
They were sent on a fishing expedition.
They went from Upstate New York all the way
to Minnesota, fishing all summer.
What they did was caught trout and caught
other fish, collected their stomach contents,
took them back to the lab, analyzed them to
see what is it that trout consume in nature.
And after they got done with all of their
analyses, it averaged out to be about 49 percent
protein, and I noted here there was no adjustment
for chitin because, you know, trout eat a
lot of insects, so that’s a little … maybe
a little high.
Fat 15 to 16 percent, fiber 8, ash 10 percent.
That was considered then the goal or the template
or what you would try to achieve in making
a feed.
Oh, jeez.
I thought I got all these stupid things out
of here.
Okay.
So, 1920s to 1940s.
Pretty much in those days, there weren’t
a lot of commercial companies; salmon and
trout feeds were made at the hatchery by the
staff.
They sourced locally available materials.
They made these wet and dry mixes that were
kind of pretty primitive, they were formed
into clumps or noodles, there was really no
sophisticated pelleting taking place, and
they were made from whatever these people
could scrounge around.
They went to slaughter houses and got animal
organs and animal parts.
Old horses, honest to God.
I met a guy who used to grind up horses.
Carp, suckers, other fish that nobody ate,
dried blends of other byproducts, dried milk,
yeast, and wheat.
Those were the kinds of ingredients that were
used.
Alright, Ron.
Oops.
Okay, Ron, let’s go back one.
So to summarize this, a lot of development
of these diets led to this type of formulation:
whitefish meal 40 percent, cottonseed meal,
wheat meal, brewer’s yeast, dried skim milk,
fish oil.
This is kind of simplifying this just for
purposes of this talk.
Having a crude protein content of 49 percent—this
would be a dried diet, fat, ash.
So, you see it kind of reflects more or less
what our boys from Portland had done in 1924.
Anyway, prior to this … or prior to the
1950s, all of these feeds that were developed
were really done by trial and error; they
were empirically derived.
We really … we did a little bit of the qualitative
dietary requirement from some nutrients but
not much of the quantitative requirements,
and what really was needed at that time was
what you’ve been talking about here, what
Frosty just discussed, and that is the need
for a purified or semi-purified diet to be
used in research.
There have been a lot of attempts to do this,
but none of them had been successful for more
than a short period; they were all nutritionally
inadequate.
But then again, what happens?
The right person came along at the right time
and moved the field forward, and that was
my professor and the guy considered the father
of fish nutrition, John Halver, and I call
him the “accidental fish nutritionist.”
He had started out to become a human nutritionist.
He was a student at the University of Washington
in a Ph.D. program, and at that time in the
early ‘50s there was a fisheries guy named
Donaldson who was just a giant in the field.
He came to the Chairman of the Department
of Biochemistry and he said, “I’ve got
a problem with our fish, our fish are anemic,
we don’t know what the deal is, we don’t
know how to cure it; it’s really a problem.”
And the Chairman of the department yelled
out the door, “Halver, get in here.”
This is what he told me.
He said, “This guy needs help; see what
you can do.”
Now, just at that time, folic acid and B12
had been recently identified and purified,
and they happened to have some.
So, Halver went down to the little hatchery,
injected these fish with folic acid and B12,
and lo and behold, in 7 to 10 days the anemia
was resolved.
Halver was pretty smart.
He thought, “Wow, these guys don’t know
anything; this is an opportunity for me.
Maybe I won’t be a human nutritionist.”
He proposed a Ph.D. project to develop a vitamin-free
test diet to study vitamins.
He did this based on casein, gelatin, and
dextrin; it worked.
He then used this diet to determine the qualitative
and then quantitative requirements of Donaldson’s
salmon.
I got ahold of his original mix sheet.
I know you can’t read that, but it looks
like a crazy recipe out of a cookbook from
the 40s or 50s: Mix this, one part of that,
put on a yellow hat, do this, do that.
But boiled down, this is what it looked like.
It had vitamin-free casein, dextrin, and gelatin;
wheat starch; CMC; alpha cell vitamin and
minerals; amino acid mixture; choline chloride.
This was a moist diet.
It was made into noodles, it was kept frozen,
and its crude protein was about 34 percent
on an as-is basis; fat, 17; ash … wait a
minute.
Maybe that’s not right.
Well anyway, that’s what it looked like.
So, what Halver did … the semi-purified
was the … you know how this works.
Make this efficient diet; make your vitamin
pre-mix; leave out the nutrients you want
to study; have graded levels—in Halver’s
case, six to eight levels; feed the fish until
they gain about three times their initial
weight, if possible—that didn’t always
happen; and then measure response variables.
Now, this is back in the 1950s here.
So, their response variables were relatively
simple: weight gain, survival, feed conversion
ratio.
A little later they developed microbial assays
with which they could analyze for vitamin
levels in livers.
A little later they got active and developed
some enzyme assays, histology.
But remember, at this time, what was it?
No HPLC, no GCs, no mass specs.
This was all done at a pretty simple level.
Dang it.
So, this is the kind of stuff you would get.
This happens to be a microbiology assay of
thiamine, and you can see the statistical
analysis was a broken line approach, and this
is the kind of data that was generated during
that time showing, in this case, a maximum
liver thiamine content at about 8 milligrams
per kilogram diet.
This diet was modified to then work to study
amino acid requirements.
In this case, vitamin-free casein and gelatin
were reduced, an amino acid mixture was added,
and away they went, leaving, of course, one
amino acid out at a time—same diet, basically.
So, this is the kind of data they got.
In this case, this is the arginine requirement.
You can see the fish start at about 2 grams
in average weight.
The dark line is the deficient diet; they
lost weight slightly over the course of the
10-week experiment.
The fish that had arginine grew at a fairly
nice rate.
They didn’t quite make the goal of three
times their initial weight, but they did grow,
and at 6 weeks Halver split his deficient
group into two, replacing arginine in one
group with the deficient diet, and you can
see here they started to grow.
That’s the kind of study that was done back
then.
So, using this approach over years of study,
the nutrient requirements were more or less
worked out.
Ten essential amino acids were identified,
omega-3 fatty acids were shown to be essential,
energy mainly supplied by lipids and protein—this
is in salmon, carbohydrates really weren’t
that good as an energy source.
Fifteen essential vitamins, minerals, carotenoids
subsequently were shown to be needed for viable
eggs, and of course minerals … there are
other minerals that are essential, but because,
as Frosty mentioned, fish live in water, a
lot of these minerals can be obtained directly
from water.
This is the NRC requirement for salmon—salmon
fingerlings, I might add—showing that it
pretty well worked out.
Nothing too exciting here, but anyway, there
they are.
They kind of formulate or list the data now
still that’s to this day is used formulate
commercial feeds.
I want to just give credit here to work done
at Oregon State University by a couple of
guys I knew: John Castell and Russ Sinnhuber.
They did very, very interesting work on omega-3
fatty acids showing that … they used 1 percent
of the diet as omega-3s, and these can be
EPA or DHA.
They also identified essential fatty acid
deficiency signs, including heart pathology,
fainting—Have you ever seen a fish faint?
Probably not—anyway, poor growth.
It took a long time to make these fish deficient.
Previous diet history had quite a great effect
on the onset of deficiency, how long it took
to get it, and then later on this requirement
was confirmed with Pacific salmon.
So, Halver assembled all this material and
published his first book in 1972.
This isn’t it; I loaned mine to a guy who
took it to Vietnam, and it was eaten by ants,
I swear to God.
So, I lost my first copy.
Anyway, this is the third copy.
Halver and I did this together in 2002, but
his first book was really a revelation.
It was the first real treatise on fish nutrition,
and it really broke the tradition in the fish
nutrition world in that it wasn’t written
from … by a bunch of fish biologists.
It was written by a biochemist, so it really
had a biochemical approach.
It established fish nutrition as a genuine
academic topic, and we are just starting to
work on the fourth edition of this book.
In ’73, a year later, up came the first
NRC bulletin on fish—that’s the green
one here—and subsequent bulletins have been
issued up to 1993 was the most recent one
up until the “the” most recent one, which
I’ll talk about in a minute—that being
this book right here.
And two of the people who worked on this who
are in the room, Del Gatlin and Lou D’Abramo
were on our committee, so thank you for that.
Thank you for your service, as they say.
We have a fish nutrition … this was published
in 2011 and is the most recent compilation
of nutrient requirements of fish and shrimp.
All of it is getting to be a little old already.
Here’s what I see as a problem you have
with zebrafish.
Here’s a copy of one of the tables from
this book showing the freshwater fish requirements
for amino acids, and you can see and compare
it in this slide the old 1993 and 2001 … or
2011 version—different colors there.
Channel catfish—we know a lot about the
amino acid requirements of catfish, we know
a lot about rainbow trout, we’re getting
to know a lot more about common carp and tilapia;
these are both very important commercial species.
Zebrafish: nothing.
We’ve got nothing.
Vitamins, same story.
You know pretty well, you can see, if you
really want to dig into this into detail,
the requirement levels did not change too
much in our new bulletin.
They’re pretty much the same, a few little
changes, little tweaks here and there.
Again, zebrafish: not much there.
Alright.
You’ve seen this already.
This is minerals.
Do I have to explain this one?
No, I think we get the picture here.
We don’t know too much.
Okay.
One of the outcomes of the development of
these data for nutrient requirements and application
of this information in the production of the
commercial diets has been a profound change
in the approximate composition of feeds for
salmon and trout—this is trout, in this
case—and you can see since the ‘70s the
level of total protein in blue has increased
from the mid-30s up to the low 40s.
But more importantly, the level of digestible
protein has increased, and the gap between
total and digestible protein has gotten smaller.
The other thing to note is that the level
of dietary fat has gone up a lot.
Currently it says here 22; this slide’s
a little out of date—people are even pushing
30 percent now in some diets.
And what’s happened in commercial production
is that this has really changed the economics
of production.
In the 1970s the food conversion ratio of
salmon and trout was 2, or even higher than
2, and over time with the improvements in
diets you can see it’s progressively decreased
such that we can know see—well, there’s
2000 … boy, I’ve got to update these slides—it’s
now down to around 1.2 for trout and about
1 percent … excuse me, 1.2 for trout and
as low as 1 for production of up to 3 kilo
salmon.
It’s a tremendous improvement from where
we were before this information was used to
formulate feeds.
So now, I’m going to switch gears a little
bit.
How many of you took sociology when you were
in college?
Everybody did.
Everybody knows what this is: Maslow’s hierarchy
of needs.
So, you know that—probably from your reading—that
the hierarchy of needs starts out very basic:
the basic requirements for survival, physiological
needs, food, water, air, shelter are the bottom
level.
Safety and security needs, social needs, friends,
family, community, recognition, and eventually
you really are lucky and good and a lot of
effort you can become self-actualized.
Well, I’ve created something called the
Hardy hierarchy of feeds.
So, I just dreamed this up here to kind of
show some of the components … some of elements
that relate to our eventual goal of producing
sustainable feeds in commercial production.
So again, we have a similar setup here.
Nutritional requirements to sustain life are
at the bottom.
As we get more sophisticated, we can refine
diets to supply nutrients needed for optimal
growth and fish health.
Feed ingredient knowledge is a component of
this; it leads to more sophisticated feed
formulations and eventually to sustainable
feeds.
So, here’s how I see it.
I think trout are up there.
We’re doing pretty well with trout.
We’re up there where we can formulate very
sophisticated feeds, and we’re almost to
the point where we can produce sustainable
feeds.
I’m sorry to tell you this: The zebrafish
are way down there.
You have diets that seem to me capable of
sustaining life, but as far as the nutritional
needs for optimal growth and health, I don’t
think you’ve gotten there yet.
With trout, we still need to learn some more
information, particularly on the nutritional
need for optimum growth and health, using
gene expression and other etiological parameters.
We’re getting a better idea of the effects
of diets and nutrition health.
Feed ingredient knowledge—we have developed
extensive databases on digestibility of nutrients
and a whole array of common feed ingredients
that really help us when we get to the next
level, which is feed formulation and understanding
better interactions among nutrients and ingredients.
And, like I said earlier, we’re almost … the
production of sustainable feeds is really
within our reach.
We can see it coming within the next 5 or
10 years.
Oh, boy.
I had maybe one too many coffees this morning.
So, with zebrafish, here’s what I see.
Nutritional needs for optimal growth and health:
not too much is known.
Feed ingredient knowledge: some is known;
you know some ingredients are good and bad.
But when you get to feed formulation and feed
ingredient interactions it’s almost impossible,
if not difficult, anyway, without knowledge
of nutritional requirements.
You just can’t do it, meaning you cannot
achieve optimized diets for whatever purpose:
medical research, physiological or developmental
studies, and so on.
So, why is this important?
I think you all realize this.
Frosty little bit alluded to this.
It’s really impossible to formulate feeds—nutritionally
balanced feeds—for zebrafish if we don’t
know what the nutritional requirements are.
You just can’t do it.
And we all know that feed ingredients affect
physiological functions.
Again, this was said earlier.
There are a couple of examples that we know
there are signaling nutrients that affect
energy allocation, protein accretion, and
so on.
We know that some of these ingredients contain
phytoestrogens.
One of the ingredients in particular, insect
hormones, are kind of an unknown but may affect
reproduction.
And as Frosty said … Frosty really said
a lot of things that I was going to say, so
I guess that’s good, but I’ll repeat it
again, and that is: Feed ingredients vary
in nutrient contents.
They vary with season, they vary with source,
they vary with raw material quality and processing
methods.
The most variable ones are fish meal, insect
meals, and land animal meals.
And fish meal, I’ll tell you, it’s an
amazing product.
It’s a very complex material that contains
all of the essential nutrients—not all of
them, but many of them that we need; high-energy
content; but it also contains other biologically
active compounds: anabolic steroids, organic
pollutants, heavy metals.
Insect meals are interesting.
Insects are like balloons.
If you feed them they basically take up whatever
you feed them, so if you feed them certain
fatty acids they’re going to accumulate
those fatty acids.
So, they’re almost like a blank canvas,
like a balloon you fill up on nutrients.
So, it’s important to know what substrates
are used to raise these insects and, of course,
what species you’re talking about.
Insect meal is not a commodity like casein
or (indiscernible) defined ingredient; it
can be a lot of different things in terms
of nutrient content.
So, zebrafish diets.
We know that there are closed formula diets
that are commercially available from feed
manufacturers.
Some are designed specifically for zebrafish,
others are designed for other species but
work okay with zebrafish.
They are kind of at the bottom level of our
hierarchy of needs.
They support decent growth and health, they
don’t kill fish off right off the bat, but
they’re not specifically designed for zebrafish.
And then (indiscernible) the zebrafish literature,
I see there are practical diets that are used
that contain unrefined ingredients such as
the ingredients I mentioned: fish meal, insect
meal, grains, (indiscernible) meals, not to
mention the starches.
And then we also have some of the purified
diets that contain highly refined ingredients:
casein, gelatin, (indiscernible) they’ve
got some nice mash diets there.
But as a fish nutritionist coming from the
commercial sector I was kind of surprised
when I looked at the literature when preparing
for this talk at some of the stuff that caught
my eye.
And again, Frosty said this—he’s absolutely
right—insufficient information on experimental
diets is a major surprise to me, anyway, in
looking at some of the published papers in
the zebrafish literature.
What we need there … we need to know feed
formulations and sources of ingredients.
We need to know chemical analysis—not every
chemical on the face of the Earth, but certainly
the ones that are important nutritionally.
And just to let you know, as an editor of
a scientific journal—I think Delbert is
an editor … I know Delbert is an editor
and he probably has the same opinion as I
do—this information is required by aquaculture
journals.
If I get a paper submitted to me that doesn’t
convey this information, it’s rejected at
the inter-trial level.
Goodbye.
We don’t even consider it.
The other thing that surprised me was the
source of commercial feeds is often not given
other than, “Oh, we bought a flake bag,”
or “We fed worms,” or we did this or we
did that.
That surprised me, too.
The other thing that surprised me was that
many of the papers that are published in this
field are published in the journal Zebrafish.
Guess what?
That’s not accessible to people like me
unless I want to shell out $51 to get a reprint.
I was surprised that that isn’t open access
or at least accessible by normal or commonly
used literature search engines, so there you
go.
So, in the interest of being politically correct,
I took this guy’s name off this paper, but
I thought I’d give an example of what not
to do.
This is a paper published in 2017—pretty
recent paper.
The point was to study carnitine.
It was done in China.
I know you probably can’t read all of this,
but what the experimenter did was he bought
a commercial zebrafish diet made in China,
he gave the source of who produced it, some
company.
That was it.
Not another word about that diet.
He then took “dough flour”—dough, he
called it and I dug around and found out that’s
wheat flour—and mixed up carnitine by hand,
dried it out, and then his experiment, which
is the lower level of these little boxes here,
were the control feed, which was 1 percent
dough and 3 percent of this mystery diet;
and then in controlled fasting, he just fed
the dough, and then he fed the dough with
carnitine and the dough … anyway, mix and
match.
Interesting study, but how do you know?
What’s missing here?
The diets weren’t analyzed for carnitine.
What’s in the Chinese commercial diet for
carnitine?
I don’t know.
It didn’t say.
It didn’t analyze it.
So, that’s something that we would probably
ding right off the bat if it was sent to my
journal.
I’ve got a good example here from Oregon
State University.
Let’s hear it for Oregon State.
I read this paper (indiscernible)—I gave
his name because he did a good job.
So, this was an example: If you want to see
the kind of information that should be in
these papers, this is the one to look at.
The authors presented complete information
on the experimental diet, the whole formulation,
the percentages of each ingredient, the sources
of each ingredient were given, the mineral
and vitamin premix are completely described,
chemical analysis was conducted, results were
presented, and he was looking in this case
at putting in tocopherol and analyzed all
the diets to show that the added levels were
pretty close to what was analyzed and found
to be present.
So, that’s what you want to do.
Well, some of the uncertainties … I mean,
there’s a common … you can figure this
out.
You know this.
That’s why we’re here.
We don’t know the nutritional requirements
of these fish.
They likely change with life history, somebody
mentioned strain or genetic variants, and
we don’t know that.
We don’t know what deficiencies are antagonist
interactions that confound results.
We commonly run into this stuff in commercial
diets into this problem, and it’s particularly
important when we’re looking at gene expression
studies.
We can find minor changes in diet composition
can alter gene expression and activation of
certain pathways.
Feed ingredients contained compounds that
can affect metabolism; I mean, obviously,
reproductive effects.
We’ve seen this in some … I actually did
a zebrafish study one time, and it changed
their reproductive status.
Redox status was mentioned earlier.
And then, another thing that was mentioned
earlier—there’s a lot of good things here
and I just want to give him credit here, Frosty
did—and that is diets can vary in nutritional
content from batch to batch.
Closed formulations may change.
Producers can make small changes in using
different substituting ingredients or altering
the level of the same ingredient—small changes—and
more importantly, and people don’t realize
this, a lot of these ingredients that are
used, not highly refined ingredients like
casein, but the natural ingredients, shall
we say, can vary a lot, and this may affect
your results.
So, going forward, best practices should be
followed.
Just to repeat what I said, zebrafish studies
should include a commercial reference diet
to facilitate comparison among labs or between
labs.
Excuse me.
I think studies should include an open formula
standard zebrafish diet; we do this in our
trout and salmon work.
The details of the feed should be provided:
like I said, formulation, source of ingredients,
the composition.
And, again, I’m going to say that this Miller
paper really impressed me; it’s a good paper.
And then, the nutritional requirements for
the three life stages should be identified;
that kind of worried me a little bit.
Zebrafish grow so fast, and in other species
of fish there are really three distinct life
history stages: baby fish, juveniles—most
juveniles—and then reproductively active
fish.
So, that’s something that needs to be investigated.
One good bit of information, a good development,
is that there are related cyprinid species
that are being grown in great numbers now
in China just over the last 10 years.
They’re grown for a food fish, they’re
bigger fish, but they’re related, and so
maybe this can be a starting point for looking
at some of the nutritional requirements for
zebrafish.
Oh, rats.
Let me back one up.
So the positives: You do have some good semi-purified
feeds.
Again, just to repeat, I think it would be
very easy … not easy, but it wouldn’t
take very long, I don’t think, to at least
identify and quantify the nutritional requirements
of zebrafish.
The interesting thing about zebrafish is you
can do studies pretty fast, short life history,
you can do 15-day feeding trials and probably
work some of these out pretty quickly.
Another advantage we have, a positive, is
that we have new response variables compared
to the early salmon days when it was basically
weight gain and survival and other easily
measured metrics.
The other thing I thought about was we kind
of know the boundaries now of some of these
nutritional requirements, so we can narrow
in more specifically to the levels that we
think are important without starting from
scratch.
And again, as I said, other cyprinids are
reared for food fish, and that will get us
off to a good start.
And I really like this workshop.
I’m excited to come here.
I’ve been thinking about this for a long
time, at least 10 or 15 years, saying, “What’s
with those zebrafish guys?”
How are they doing this work?
I think the time has come that zebrafish nutritionists
can really benefit from the accumulated knowledge
and experiences that we’ve developed in
the other sector of aquaculture, meaning food
fish production, so I think you can jumpstart
this and get at it pretty quickly.
So, with that I’ll conclude and say, just
to repeat with my little triangle there that
I showed—my pyramid—I think you’re now
at a similar stage that trout and salmon were
in the 1950s, but I don’t think you’re
going to have to wait 50-60 years to get to
where we are today.
The new knowledge and the new connections
we have, with a few years of effort, we can
catch up and clean things up here pretty fast.
So, thanks very much.
STEPHEN WATTS: Thank you very much, Ron.
He pulls no punches.
Any comments?
RONALD HARDY: I’m old enough I don’t have
to.
STEPHEN WATTS: There we go.
That’s right.
Any comments?
Yes?
UNIDENTIFIED MALE: Yeah, I was glad to hear
you say that if you’ve got a paper that
really did a poor job of describing the fish
diet that you would reject outright, and I
think it’s worth saying again.
I know a lot of people have already said it,
but I think that’s one of the problems in
general, certainly in the rodent literature,
and hopefully we can avoid this with the zebrafish
literature, which is papers are being submitted
and reviewed by people that don’t necessarily
have the nutrition background to review them,
and so they’ll get accepted, and they’ll
compare a cereal base to a purified or something
that’s—as we’re agreeing here—is fairly
inappropriate, and it will get published in
a very top-tiered journal, and that kind of
feeds forward.
And I’m also wondering if this can get backed
up to the grant review stage where, you know,
folks writing these grants … this stuff
can get caught at that stage—this whole
rigor and reproducibility thing.
We shouldn’t really be funding poorly designed
studies, especially diet-wise.
So, to me that’s just a constant thing you
see from a nutritionist perspective in the
literature—bad diet upon bad diet upon bad
diet—and it just feeds forward, because
then that paper gets cited as the gospel,
and now we have a problem because we’re
citing bad data.
RONALD HARDY: That’s easy to fix.
At the review stage, if NIH were to reach
out a little more broadly into the commercial
or our world, basically, we could provide
that kind of input.
We don’t know the details of some of the
work that you’re doing, but we can certainly
provide input on that.
The other thing I would say, I would like
to see more cross-pollination and less stovepiping
between aquaculture, nutritionists, like Delbert
and Lou and myself, and the zebrafish community,
because I think both of us could profit from
that.
UNIDENTIFIED MALE: So, just a quick comment
about the journal Zebrafish and about your
statement.
Steve Ekker, the editor of Zebrafish, has
made an effort to bring these kinds of topics
more into the consciousness of the zebrafish
community, and Zebrafish journal was a vehicle
that was best suited for that, but unfortunately
it’s not available to everyone because it’s
not open-source unless the authors can afford
the fee.
But nevertheless, I think the starting point
has been given to make people aware that nutrition
is something we have to pay attention to this,
and I would be very grateful if the cross-pollination
aspect that you mentioned would really happen
and we can, you know, successfully publish
zebrafish papers in nutrition journals and,
you know, make a contribution to developing
better knowledge about this issue.
RONALD HARDY: Well, I … just a comment that
I do see these papers in Aquatic Toxicology
and journals like that, so there is some … every
paper isn’t published in the Zebrafish journal.
There are others … there’s some that get
out.
UNIDENTIFIED MALE: Just to provide some background,
I think one key point is that zebrafish sort
of came from or evolved out of an interest
in studying early development, and so the
thing that was only optimized was, do the
fish grow quickly, and how many eggs do I
get?
And although … and so that community started,
but then the pressure to address so many aspects
of the physiology emerged such that the whole
field now is asking questions about cardiovascular
disease and heart development and regeneration
and a ton of physiology building on essentially
no knowledge beyond this anecdotal, like,
okay, you can raise fish this way, that way,
or another way.
So, I think we never had this period of optimizing
growth the way a commercial field would have,
because you need an adult fish at the end,
and the zebrafish for a while didn’t care
about anything beyond, like, 48 hours, and
even that was late.
RONALD HARDY: That’s a good point.
UNIDENTIFIED MALE: So, I think we are … but
that said, the whole community is changing
because really interesting physiology is there
to be discovered with this model.
So, I think we just have to do … somewhere
we have to figure out a funding stream to
really do these.
Really, we could even start with your tables.
Like you said, it shouldn’t take us 20 years,
because we could start with what aquaculture
has done and just see how close does the fish
match that?
RONALD HARDY: That’s a fair point.
You’re right.
It’s evolved from its origins to much broader.
UNIDENTIFIED MALE: And a lot of editors in
developmental biology probably know nothing
or very little about this concern, because
if you produce an egg and it develops in 24
hours, it’s fine.
RONALD HARDY: Yeah.
UNIDENTIFIED MALE: A comment about what you
were saying earlier.
Let’s just make it clear: We don’t know
what a bad diet is, so we cannot regulate
any of that at this point because, as Dr.
Hardy said, we just don’t know a whole lot
about nutrition and what are in these diets
and how it affects the zebrafish.
So, we can’t regulate from the front end
saying, you know, “We’re not going to
approve your study because you don’t have
a good diet,” but we can say, you know,
“When you do publish this, publish some
nutritional information with it,” okay?
And that can be as simple as, you know, “We’re
using a flake diet from the pet store.
Let’s send it in to get analyzed for a quick
…” Even if it’s a proximate analysis,
right?
That would be a good start to get the protein,
the lipids, and so on and so forth, because
we don’t even do that at this point.
Eventually, we’d like to improve that and
get even more rigorous with some of those
reports, but start out very basic until we
actually find out what we know is good and
bad for zebrafish.
So, that may be a matter of semantics but
let’s try not to … when we talk about
this to our fellow colleagues when we leave
here, let’s not talk about those types of
things because that’s not really what we’re
doing.
We don’t really know what’s good and bad
at this point.
STEPHEN WATTS: We’ll go ahead and move on
and we’ll have additional discussion later.
Thank you very much, Ron.
RONALD HARDY: Thank you.