Does Breeding System Contribute to Rarity of Goldenseal (Hydrastis canadensis)?


Sanders, Suzanne

The American Midland Naturalist 152(1): 42

2004


Goldenseal (Hydrastis canadensis L.) is an herbaceous perennial species that is becoming more rare within its range. Hydrastis canadensis populations are highly isolated and pollen flow between these populations may be restricted. I examined the breeding system of H. canadensis to determine if it may be limiting seed set due to increasing isolation of individuals and populations from one another. I tested fruit set in treatments designed to detect the presence of apomixis, passive autogamy, active autogamy, short distance outcrossing and long distance outcrossing. No fruit set occurred in flowers that were emasculated and bagged, suggesting the species is incapable of apomixis. However, low rates of fruit set were found in all other treatments, suggesting a mixed mating system in which both selfing and outcrossing may occur. Pollen transfer between highly isolated populations can result in fruit set, as can within-population pollen transfer. The breeding system type appears unlikely to be a major factor limiting the distribution or abundance of H. canadensis. However, overall low rates of fruit set may be important demographically. Reprinted by permission of the publisher.

Am.
Midl.
Nat.
152:37-42
Does
Breeding
System
Contribute
to
Rarity
of
Goldenseal
(Hydrastis
canadensis)?
SUZANNE
SANDERS
Department
of
Biology,
West
Virginia
University,
P.O.
Box
6057,
Morgantown
26506
ABSTRACT.—Goldenseal
(Hydrastis
canadensis
L.)
is
an
herbaceous
perennial
species
that
is
becoming
more
rare
within
its
range.
Hydrastis
canadensis
populations
are
highly
isolated
and
pollen
flow
between
these
populations
may
be
restricted.
I
examined
the
breeding
system
of
H.
canadensis
to
determine
if
it
may
be
limiting
seed
set
due
to
increasing
isolation
of
individuals
and
populations
from
one
another.
I
tested
fruit
set
in
treatments
designed
to
detect
the
presence
of
apomixis,
passive
autogamy,
active
autogamy,
short
distance
outcrossing
and
long
distance
outcrossing.
No
fruit
set
occurred
in
flowers
that
were
emasculated
and
bagged,
suggesting
the
species
is
incapable
of
apomixis.
However,
low
rates
of
fruit
set
were
found
in
all
other
treatments,
suggesting
a
mixed
mating
system
in
which
both
selfing
and
outcrossing
may
occur.
Pollen
transfer
between
highly
isolated
populations
can
result
in
fruit
set,
as
can
within-population
pollen
transfer.
The
breeding
system
type
appears
unlikely
to
be
a
major
factor
limiting
the
distribution
or
abundance
of
H.
canadensis.
However,
overall
low
rates
of
fruit
set
may
be
important
demographically.
INTRODUCTION
Goldenseal,
Hydrastis
canadensis
L.,
is
a
long-lived
herbaceous
perennial
plant
of
the
eastern
deciduous
forest.
Recent
investigations
have
shown
that
this
species
is
in
decline;
previously
documented
populations
have
either
become
reduced
in
size
or
extirpated
(Sinclair
and
Gaffing,
2000a;
Sanders
and
McGraw,
2002;
Mulligan
and
Gorchov,
2003).
Suggested
causes
of
this
decline
include
harvest
(Robbins,
2000;
Mulligan
and
Gorchov,
2003),
stochastic
events
(Sanders
and
McGraw,
2002),
succession
(Sanders
and
McGraw,
2002)
and
changes
in
disturbance
modes
and
patterns
(Sinclair
and
Catling,
2000b).
Species
that
have
experienced
some
level
of
decline
may
be
further
susceptible
to
other
stresses,
leading
to
a
feedback
loop.
One
such
feedback
loop
may
exist
between
species
abundance
and
breeding
system
characteristics.
Breeding
systems
are
most
likely
to
limit
seed
set
if
a
species
obligately
or
preferentially
outcrosses
and/or
available
mates
are
spatially
separated
across
the
landscape
(Demauro,
1993;
Weekley
and
Race,
2001).
Mate
limitation
resulting
from
obligate
outcrossing
has
been
associated
with
rarity
in
numerous
studies
(Evans
et
al.,
2003;
Messmore
and
Knox,
1997).
In
clonal
plant
species,
only
one
or
a
few
genets
may
be
present
within
a
given
patch.
Thus,
for
clonal
species
with
an
obligate
outcrossing
requirement,
viable
seed
set
may
necessitate
pollen
flow
between
patches
(Wilcock,
2002).
Hydrastis
canadensis
is
a
clonal
species
that
may
be
adversely
affected
by
breeding
system
characteristics.
This
species
forms
dense
patches
of
a
few
to
greater
than
1000
ramets.
These
patches
are
frequently
sparsely
distributed
across
the
landscape,
such
that
many
patches
are
isolated
from
others
by
great
distances
(McGraw
et
al.,
2003).
While
the
degree
of
genetic
variation
within
patches
is
unknown,
the
extensive
clonal
growth
pattern
suggests
that
only
one
or
a
few
genotypes
may
be
present
within
a
given
patch.
Thus,
if
H.
canadensis
is
an
obligate
outcrossing
species,
seed
production
and
viability
may
be
hindered
due
to
mate
limitation.
Preliminary
studies
conducted
at
two
Ontario,
Canada
sites
indicate
H.
canadensis
may
be
self-compatible
(Sinclair
et
al.,
2000).
However,
the
authors
used
only
three
plants
to
infer
37
38
THE
AMERICAN
MIDLAND
NATURALIST
152(1)
this.
While
this
suggests
that
autogamy
occurs,
the
degree
of
passive
vs.
active
autogamy,
and
the
relative
contribution
of
selfing
and
outcrossing
to
reproductive
success
remains
unknown.
The
objective
of
the
present
study
was
to
determine
the
breeding
system
of
H.
canadensis.
I
performed
crosses
to
test
for
apomixis,
passive
autogamy,
active
autogamy
and
outcrossing,
both
within
and
between
populations.
METHODS
Plant
material
from
three
natural
H.
canadensis
sources
in
north
central
West
Virginia
was
used:
Cheat
Canyon,
Morgantown
and
Jane
Lew.
These
populations
were
relatively
large
and
separated
by
20-120
km.
At
each
site,
stems
were
selected
for
study
during
August
2001.
Stems
selected
for
study
were
all
reproductive
in
2001
and
an
assumption
was
made
that
stems
reproductive
in
one
year
were
likely
to
be
reproductive
the
following
year.
I
based
the
number
of
rhizomes
collected
on
the
total
number
of
stems
at
each
site
that
were
reproductive
in
2001;
because
I
did
not
want
to
deplete
the
patches
of
reproductive
ramets,
I
selected
approximately
half
of
those
that
were
flowering
in
2001.
The
total
number
of
rhizomes
removed
at
each
site
was
130
from
Cheat
Canyon,
80
from
Jane
Lew
and
60
from
Morgantown.
A
numbered
aluminum
nail
was
placed
in
the
ground
immediately
uphill
of
these
stems.
A
map
was
drawn
so
that
general
locations
of
stems,
and
thus
rhizomes,
were
known.
Rhizomes
were
relocated
prior
to
emergence
on
30
March,
1
April
and
4
April
2002
with
the
map
and
a
metal
detector.
Rhizomes
were
removed,
labeled
and
replanted
in
plastic
pots
(6
cm
diam,
18
cm
deep)
in
the
West
Virginia
University
Plant
Science
Greenhouse.
Shoots
emerged
during
the
second
week
of
April,
whereupon
shade
cloth
was
placed
over
the
pots.
To
determine
breeding
system
type,
five
treatments
were
implemented:
(1)
emasculation
followed
by
bagging
(test
for
apomixis),
(2)
bagging
with
no
hand
transfer
of
pollen
(test
for
passive
autogamy),
(3)
bagging
with
hand
transfer
of
pollen
within
flowers
(test
for
active
autogamy),
(4)
emasculation
plus
within-source
pollen
transfer
followed
by
bagging
(test
for
short
distance
outcrossing)
and
(5)
emasculation
plus
between
source
pollen
transfer
followed
by
bagging
(test
for
long-distance
outcrossing).
Crosses
were
made
so
that
there
were
at
least
20
replicates
in
each
treatment
(Table
1).
Given
that
each
reproductive
ramet
produces
a
single
flower,
there
was
a
minimum
of
20
flowers
per
treatment.
Because
not
all
ramets
flowered,
the
total
number
of
ramets
in
the
greenhouse
treatments
(141)
was
less
than
the
number
collected
(270).
Crosses
were
made
so
that
the
total
number
of
flowers
in
each
treatment
would
be
approximately
equal.
However,
because
a
primary
interest
was
to
determine
if
H.
canadensis
has
the
ability
to
self-fertilize,
I
allotted
more
ramets
in
the
test
for
active
autogamy.
Crosses
were
initiated
on
12
April
2002
and
all
treatments
were
completed
by
19
April
2002.
All
emasculation
was
performed
by
clipping
anthers
off
filaments
with
scissors
prior
to
anther
dehiscence.
Pollen
was
transferred
by
removing
at
least
three
stamens
with
tweezers
and
brushing
the
anthers
over
the
desired
stigma.
Flowers
were
selected
as
a
pollen
source
when
at
least
one
anther
had
a
tinge
of
brown,
indicating
dehiscence
had
begun.
Crosses
between
a
pollen
source
and
the
designated
treated
flower
were
then
made
on
consecutive
days
until
all
remaining
anthers
on
the
source
flower
were
clearly
no
longer
viable.
In
most
instances,
crosses
between
a
given
pollen
source
and
a
designated
treated
flower
lasted
over
a
3
d
period.
Flowers
were
bagged
using
a
fine
mesh
cut
into
disks
of
approximately
10
cm
diameter
and
threaded
around
the
perimeter
with
embroidery
floss.
The
floss
was
tied
around
the
pedicel,
tightly
enough
that
it
would
not
easily
come
off,
but not
so
tight
as
to
be
constricting.
Ramets
were
considered
to
have
set
seed
when
at
least
one
pistil
of
the
flower
developed
into
a
fruit.
Seeds
were
collected
in
July
as
they
matured
and
tested
for
viability
2004
SANDERS:
GOLDENSEAL
RARITY
39
TABLE
1.—Fruit
set
percentage
for
each
treatment.
The
number
of
flowers
represented
from
each
site
is
shown
indented
and
in
parentheses
Treatment
Number
of
flowers
in
treatment
Number
of
flowers
setting
fruit
Fruit
set
percentage
1.
Emasculation
25
0 0
Cheat
Canyon
(
3
)
(0) (0)
Jane
Lew
(8)
(0) (0)
Morgantown
(14)
(0) (0)
2.
Passive
25
6
24.0
Cheat
Canyon
(8)
(2)
(25.0)
Jane
Lew
(7)
(0) (0)
Morgantown
(10)
(4)
(40.0)
3.
Active
45
8
17.8
Cheat
Canyon
(14)
(6)
(42.9)
Jane
Lew
(21)
(1)
(4.8)
Morgantown
(10)
(1)
(10)
4.
Within
source
22
3
13.6
Cheat
Canyon
(
7
)
(1)
(14.3)
Jane
Lew
(7)
(0)
(0)
Morgantown
(8)
(2)
(25.0)
5.
Between
sources
24
3
12.5
Cheat
Canyon
(6)
(2)
(33.3)
Jane
Lew
(4)
(1)
(25.0)
Morgantown
(14)
(0) (0)
6.
Field
sites
831
316
38.0
Cheat
Canyon
(106)
(21)
(19.8)
Jane
Lew
(544)
(165)
(30.3)
Morgantown
(181)
(130)
(71.8)
using
the
tetrazolium
test
(Baskin
and
Baskin,
1998).
Hydrastis
canadensis
seeds
exhibit
morphophysiological
dormancy.
After
fruit
maturity
on
the
parent
plant,
seeds
require
a
period
of
time
during
which
the
embryo
enlarges.
During
this
time,
H.
canadensis
seed
requires
exposure
to
a
warm
period
followed
by
a
cold
period
(Baskin
and
Baskin,
1998).
Additionally,
seed
germination
percentage
is
low
and
more
than
1
y
is
often
required
to
observe
maximum
germination
rates
(Davis
and
McCoy,
2000).
Because
of
this,
I
believe
the
tetrazolium
test
was
a
more
sensitive
indicator
of
crossing
success
than
seed
germination.
Differences
among
treatments
were
tested
using
contingency
analysis
(SAS
JMP,
V.5.0;
SAS,
Inc.,
2002).
The
independent
factor
was
treatment
and
the
response
variable
was
fruit
set.
Apomixis
was
determined
by
whether
or
not
fruit
set
occurred
in
emasculated
bagged
ramets
(treatment
1).
To
test
for
self
fertilization,
I
observed
whether
fruit
set
occurred
in
treatments
2
(passive
autogamy)
and
3
(active
autogamy).
The
difference
in
fruit
set
between
treatments
2
and
3
was
used
to
determine
whether
pollinators
could
potentially
increase
fruit
set.
To
test
for
ability
to
outcross,
I
observed
whether
fruit
set
occurred
in
treatments
4
(within
source
pollen
transfer)
or
5
(between
source
pollen
transfer).
To
test
if
there
is
a
preferred
outcross
mating
distance,
I
compared
treatments
4
and
5.
Finally,
I
wanted
to
determine
if
H.
canadensis
exhibits
a
preference
for
selfing
vs.
outcrossing.
I
contrasted
fruit
set
in
the
two
40
THE
AMERICAN
MIDLAND
NATURALIST
152(1)
pooled
selfing
treatments
(2
and
3)
with
fruit
set
in
the
two
pooled
outcrossing
treatments
(4
and
5).
For
all
analyses,
significance
was
determined
using
a
log
likelihood
(G)
test
(Sokal
and
Rohlf,
1995).
In
addition
to
the
greenhouse
study,
I
also
noted
fruit
set
in
the
field.
In
late
June
2002,
in
each
of
the
three
populations
from
which
the
greenhouse
study
rhizomes
were
collected,
I
counted
the
total
number
of
reproductive
(two-
and
three-leaved)
ramets
and
noted
how
many
of
these
set
fruit.
I
calculated
the
overall
fruit
set
percentage
in
the
field,
which
allowed
me
to
determine
if
fruit
set
in
the
field
was
comparable
to
that
in
the
greenhouse.
I
also
calculated
the
individual
fruit
set
percentages
at
each
of
the
three
populations
where
collections
were
made.
This
allowed
me
to
test
for
natural
variation
in
fruit
set
among
populations
using
a
log
likelihood
test.
For
all
statistical
comparisons,
a
=
0.05
was
the
threshold
level
at
which
tests
were
considered
significant.
RESULTS
Overall,
there
was
significant
variation
in
fruit
set
among
treatments
in
the
greenhouse
(contingency
analysis,
P
<
0.0001).
Fruit
set
occurred
in
all
treatments
except
treatment
1
(the
test
for
apomixis,
Table
1),
in
which
none
of
25
ramets
set
fruit.
Given
the
fruit
set
rate
in
the
field
(38.0%;
see
below),
the
probability
of
obtaining
zero
of
25
ramets
setting
fruit
is
<0.0001.
In
the
test
for
passive
autogamy
(treatment
2),
6
of
25
ramets
(24.0%)
set
fruit.
In
the
test
for
active
autogamy
(treatment
3),
8
of
45
ramets
(17.8%)
set
fruit
(Table
1).
Fruit
set
of
treatment
2
did
not
differ
from
that
of
treatment
3
(P
=
0.5342).
In
the
tests
for
ability
to
outcross,
3
of
22
within
source
crosses
(13.6%,
treatment
4)
set
fruit
and
3
of
24
between
source
crosses
(12.5%,
treatment
5)
set
fruit.
These
two
treatments
did
not
differ
(P
=
0.9090).
Fruit
set
of
the
two
selfing
treatments
(2
and
3)
did
not
differ
from
that
of
the
two
outcrossing
treatments
(4
and
5)
(P
=
0.3319),
showing
that
H.
canadensis
does
not
exhibit
a
preference
for
selfing
vs.
outcrossing.
All
seeds
produced
in
the
greenhouse
study
were
shown
to
be
viable
using
the
tetrazolium
test.
Fruit
set
in
the
three
field
populations,
ranging
from
19.1-71.8%
(mean
=38.0%;
Table
1),
differed
significantly
among
sites
(P
=
<0.0001).
DISCUSSION
An
overall
goal
of
the
current
research
was
to
assess
the
role
of
the
breeding
system
of
H.
canadensis
as
a
causal
agent
of
its
rarity.
The
rarity
of
H.
canadensis
is
atypical
in
that
this
species
occupies
a
broad
geographical
range,
but
within
this
range
local
population
size
is
small
and
niche
breadth
appears
narrow
(McGraw
et
al.,
2003).
In
ecology,
there
is
a
general
association
between
species
abundance
and
range
size,
whereby
species
with
broad
geographical
ranges
tend
to
be
both
locally
abundant
and
evenly
distributed
within
that
range
while
species
with
small
geographical
ranges
tend
to
be
very
habitat
specific,
thereby
having
both
limited
distribution
and
abundance
(Rabinowitz,
1981;
Johnson,
1998).
Hydrastis
canadensis
does
not
follow
this
relationship
between
range
size
and
species
abundance.
The
range
of
H.
canadensis
extends
eastward
from
Missouri
and
northern
Arkansas
to
the
Appalachian
Mountains,
as
far
south
as
Tennessee
and
parts
of
the
neighboring
states
and
north
to
New
York
state
and
southern
Ontario,
Canada
(Small
and
Catling,
1999).
Within
this
range,
H.
canadensis
grows
clonally,
typically
in
dense
patches,
although
dispersed
patches
and
isolated
individuals
are
occasionally
observed.
It
is
not
clear
why
H.
canadensis
is
restricted
to
the
areas
where
these
patches
are
located.
However,
because
H.
canadensis
possesses
a
mixed
breeding
system,
it
would
appear
that
the
distances
separating
populations
are
not
adversely
affecting
the
ability
to
set
fruit.
Within
a
population,
fruit
set
can
occur
via
pollen
transfer
either
between
two
2004
SANDERS:
GOLDENSEAL
RARITY
41
ramets
(which,
due
to
the
clonal
nature,
may
or
may
not
represent
two
distinct
genotypes)
or
within
a
ramet.
Additionally,
fruit
set
via
pollen
transfer
between
populations
is
possible.
Despite
the
mixed
breeding
system,
there
were
marked
differences
in
fruit
set
in
the
field
between
the
three
source
populations.
At
two
of
these
populations,
Cheat
Canyon
and
Jane
Lew,
fruit
set
in
the
field
(19.8%
and
30.3%,
respectively)
was
comparable
to
that
observed
in
the
greenhouse.
At
the
Morgantown
population,
however,
fruit
set
was
71.8%.
One
factor
that
may
influence
site
dependent
reproductive
success
is
light
availability
(Kato
and
Hiura,
1999).
The
Morgantown
site
was
on
a
south
facing
slope
with
a
canopy
somewhat
more
open
than
that
of
the
other
two
sites.
The
greater
light
availability
here
could
have
facilitated
photosynthesis,
in
turn
promoting
carbon
gain
and
fruit
set.
Increased
reproductive
success
associated
with
light
has
been
reported
in
other
forest
understory
species.
Niesenbaum
(1993)
found
that
light
availability,
but not
pollen
limitation,
influenced
fruit
set
of
Lindera
benzoin
(L.)
Blume,
an
understory
shrub
often
associated
with
H.
canadensis.
Devlin
(1988)
reported
reduced
mean
seed
number
of
Lobelia
cardinalis
plants
subjected
to
10%
of
available
light
compared
with
those
receiving
27%
of
available
light.
In
addition
to
light,
another
factor
that
may
influence
reproductive
success
of
a
population
is
the
degree
of
genetic
variation
present
(Ehlers,
1999;
Schmidt
and
Jensen,
2000).
Unfortunately,
the
degree
of
genetic
variation
present
in
H.
canadensis
patches
is
currently
unknown.
Due
to
the
extensive
clonal
nature
of
this
species,
however,
it
is
unlikely
that
there
are
a
large
number
of
genotypes
present
within
any
given
H.
canadensis
patch.
Collectively,
factors
contributing
to
low
fruit
set
at
some
sites
may
have
important
demographic
consequences
for
this
species.
There
is
evidence
that
H.
canadensis
has
high
habitat
specificity
(McGraw
et
al.,
2003)
and
recruitment
by
seed
appears
to
be
relatively
uncommon
(Sanders
and
McGraw,
2002).
Species
with
specific
habitat
requirements,
such
as
H.
canadensis,
may
require
greater
seed
production
to
increase
the
likelihood
of
deposition
into
suitable
habitat,
where
germination
may
occur
and
new
patches
may
establish.
While
we
do
not
yet
fully
understand
the
rarity
of
H.
canadensis,
the
results
of
this
study
can
be
directly
applied
in
the
management
of
this
species.
Because
fruit
set
can
occur
autogamously,
protection
of
small
isolated
populations
may
be
as
important
as
the
protection
of
larger
metapopulations.
Likewise,
this
research
advances
our
knowledge
of
H.
canadensis
conservation.
If
its
rarity
continues
to
increase,
population
establishment
in
protected
areas
may
be
a
viable
method
of
species
protection.
Where
source
material
is
limited,
population
establishment
via
a
single
source
may
be
acceptable.
Acknowledgments.
—I
thank
Sue
Myers
and
the
staff
of
the
Plant
Science
Greenhouse
for
allowing
me
to
conduct
this
study
at
their
facility
and
for
helping
construct
the
shadehouse.
I
am
also
grateful
to
Mary
Ann
Furedi
for
her
assistance
in
the
relocation
and
removal
of
rhizomes
from
the
field.
Finally,
I
thank
Dr.
James
McGraw
and
two
anonymous
reviewers
for
their
assistance
in
preparing
this
manuscript.
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SUBMITTED
30
OCTOBER
2003
ACCEPTED
8
JANUARY
2004