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Propensity of Tampons and Barrier Contraceptives to Amplify Staphylococcus aureusToxic Shock Syndrome Toxin-I

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Abstract and Figures

Although the incidence of reported cases of toxic shock syndrome (TSS) has declined in recent years, the disease continues to occur in menstruating women using the newer, less-absorbent tampons or barrier contraceptives. Extant tampons and other vaginal devices were tested for the ability to induce TSS toxin-1 (TSST-1) by a TSS strain of Staphylococcus aureus MN8, a known high-toxin producer. Tested for the first time were 20 varieties of tampons, including 2 all-cotton brands newly introduced in the United States, a polyurethane contraceptive sponge, a latex diaphragm, and a polymer menstrual collection cup. All products were washed in sterile distilled water prior to use to reduce the effect of leachable chemicals. Duplicate experiments with unwashed products were also performed. Entire tampons and other test products were immersed in brain heart infusion broth plus yeast extract (BHIY) and inoculated with S. aureus MN8, a known TSST-1 producer. After incubation, the culture supernatants were assayed for TSST-1 by gel immunodiffusion. Except for all-cotton tampons, greater amounts of TSST-1 were detected in the supernatant fluid of washed tampons than detected in those which were not washed. While TSST-1 levels in unwashed non-cotton tampons ranged from 0.5 to 8 microg/ml, when these products were washed, TSST-1 levels increased to 2-32 microg/ml. In all-cotton tampons, whether washed or not, there was no detectable TSST-1. The propensity for all-cotton tampons not to amplify TSST-1 in vitro suggests they would lower the risk for tampon-associated TSS.
Content may be subject to copyright.
Infectious
Diseases
in
Obstetrics
and
Gynecology
2:140-145
(1994)
(C)
1994
Wiley-Liss,
Inc.
Propensity
of
Tampons
and
Barrier
Contraceptives
to
Amplify
Staphylococcus
aureus
Toxic
Shock
Syndrome
Toxin-I
Philip
M.
Tierno,
Jr.,
and
Bruce
A.
Hanna
Departments
of
Microbiology
and
Pathology,
New
York
University
School
of
Medicine,
New
York
University
Medical
Center,
New
York,
NY
ABSTRACT
Objective:
Although
the
incidence
of
reported
cases
of
toxic
shock
syndrome
(TSS)
has
declined
in
recent
years,
the
disease
continues
to
occur
in
menstruating
women
using
the
newer,
less-absorbent
tampons
or
barrier
contraceptives.
Extant
tampons
and
other
vaginal
devices
were
tested
for
the
ability
to
induce
TSS
toxin-1
(TSST-1)
by
a
TSS
strain
of
Staphylococcus
aureus
MN8,
a
known
high-toxin
producer.
Tested
for
the
first
time
were
20
varieties
of
tampons,
including
2
all-cotton
brands
newly
introduced
in
the
United
States,
a
polyurethane
contraceptive
sponge,
a
latex
dia-
phragm,
and
a
polymer
menstrual
collection
cup.
Methods:
All
products
were
washed
in
sterile
distilled
water
prior
to
use
to
reduce
the
effect
of
leachable
chemicals.
Duplicate
experiments
with
unwashed
products
were
also
performed.
Entire
tampons
and
other
test
products
were
immersed
in
brain
heart
infusion
broth
plus
yeast
extract
(BHIY)
and
inoculated
with
S.
aureus
MN8,
a
known
TSST-1
producer.
After
incubation,
the
culture
supernatants
were
assayed
for
TSST-1
by
gel
immunodiffusion.
Results:
Except
for
all-cotton
tampons,
greater
amounts
ofTSST-1
were
detected
in
the
superna-
tant
fluid
of
washed
tampons
than
detected
in
those
which
were
not
washed.
While
TSST-1
levels
in
unwashed
non-cotton
tampons
ranged
from
0.5
to
8
pg/ml,
when
these
products
were
washed,
TSST-1
levels
increased
to
2-32
g/ml.
In
all-cotton
tampons,
whether
washed
or
not,
there
was
no
detectable
TSST-1.
Conclusions:
The
propensity
for
all-cotton
tampons
not
to
amplify
TSST-1
in
vitro
suggests
they
would
lower
the
risk
for
tampon-associated
TSS.
(C)
1994
Wiley-Liss,
Inc.
KEY
WORDS
TSS,
toxins,
menstrual
he
incidence
of
reported
cases
of
menstrual
toxic
shock
syndrome
(TSS)
that
fulfill
the
strict
ep-
idemiologic
case
definition
criteria
has
declined
since
high-absorbency
fibers
such
as
polyacrylate
rayon,
polyester,
and
carboxymethylcellulose
were
removed
from
tampons.
Nonetheless,
the
disease
continues
to
occur,
especially
in
young
menstruat-
ing
women
using
the
newer,
less-absorbent
tam-
pons
made
of
viscose
rayon
with
and
without
cot-
ton.
Women
using
barrier
contraceptives
are
also
at
increased
risk
for
TSS.
Whole
tampons
or
their
components
have
been
reported
to
amplify
the
pro-
duction
of
TSS
toxin-
(TSST-
1)
by
TSS
strains
of
2--8
Staphylococcus
aureus
grown
in
their
presence.
Until
recently,
however,
all-cotton
tampons
were
not
available
in
the
United
States
for
either
analysis
or
use.
A
comparison
of
the
propensity
for
these
newer
tampon
products,
including
the
newly
avail-
Address
correspondence/reprint
requests
to
Dr.
Bruce
A.
Hanna,
Bellevue
4W1,
NYU
Medical
Center,
560
First
Ave.,
New
York,
NY
10016.
Basic
Science
Article
Received
May
13,
1994
Accepted
July
21,
1994
TAMPONS,
BARRIER
CONTRACEPTIVES,
AND
TSST-1
TIERNO
AND
HANNA
able
all-cotton
tampons
as
well
as
other
vaginal
devices,
to
amplify
TSST-1
production
is
the
sub-
ject
of
this
report.
MATERIALS
AND
METHODS
One
hundred
sixty-four
extant
tampons
and
ex-
tinct
control
tampon
representing
20
varieties,
6
Today
contraceptive
sponges,
6
Tassaway
menstrual
collection
cups,
and
40rtho
All-Flex
diaphragms
were
assayed
for
the
ability
to
induce
TSST-1
by
S.
aureus
MN8,
a
known
high-toxin
producer.
Prior
to
the
assay,
all
products
were
soaked
overnight
in
100
ml
of
sterile
distilled
water
at
room
tempera-
ture
in
order
to
reduce
the
leachable
chemicals
con-
tained
within
the
products.
After
washing,
whole
tampons
and
sponges
were
aseptically
removed,
squeezed,
then
washed
again
in
sterile
distilled
wa-
ter
for
1-2
h.
Whole
products
were
squeezed
again,
then
immersed
into
40
ml
of
brain
heart
infusion
broth
with
1%
yeast
extract
(BHIY)
which
con-
tained
approximately
l0
s
cfu
(colony
forming
units)/ml
of
S.
aureus
MN8,
a
known
high-toxin
producer
originally
recovered
from
a
case
of
TSS.
Since
menstrual
cups
and
diaphragms
are
made
of
non-absorbent
material,
they
were
not
squeezed
af-
ter
washings.
A
duplicate
set
of
experiments
with
unwashed
products
was
also
performed
as
were
ex-
periments
with
BHIY
controls.
All
experiments
were
incubated
overnight
in
an
atmosphere
of
5%
COg
at
37C
without
shaking,
then
assayed
for
TSST-1
by
a
gel
immunodiffusion
procedure
pre-
viously
described.
2
Final
bacterial
populations
were
determined
by
a
standard
viable
plate
count.
The
Mann-Whitney
test
was
used
to
compare
all
sets
of
3
or
more
data
points
with
still
controls;
the
null
hypothesis
was
rejected
with
P
<
0.01.
The
antic-
ipated
conclusion
is
that
tampons
(except
cotton
varieties),
diaphragms,
and
sponges
affect
toxin
production.
RESULTS
The
composition
and
relative
absorbency
of
the
vaginal
devices
tested
are
listed
in
Table
1.
The
results
of
experiments
to
detect
TSST-1
in
the
su-
pernatant
of
S.
aureus
cultivated
in
the
presence
of
washed
and
unwashed
products
as
well
as
controls
are
listed
in
Table
2.
The
recorded
ranges
of
TSST-1
are
an
average
for
each
product
style
and
type.
These
data
indicate
that
the
level
of
toxin
production
was
dependent
upon
the
product
tested.
In
general,
except
for
all-cotton
tampons,
greater
amounts
of
TSST-1
were
detected
in
the
superna-
rant
fluid
of
washed
tampons
than
detected
in
those
that
were
not
washed.
While
TSST-1
levels
in
unwashed
tampons
ranged
from
0.5
to
8
Ixg/ml,
when
these
products
were
washed,
TSST-1
levels
increased
to
2-32
Ixg/ml.
The
greatest
stimulation
of
TSST-1,
by
extant
brand
tampon
types,
was
observed
with
OB
tampons
(9.3-18.7
Ixg/ml),
Ko-
tex
(6.7-13.3
Ixg/ml),
Playtex
(6.7-9.3
Ixg/ml),
Tampax
(4-8.0
txg/ml),
and
CO-OP
(3.3-6.7
txg/
ml).
The
2
all-cotton
tampon
brands,
Terra
Femme
and
Natracare,
produced
no
measurable
toxin
whether
washed
or
unwashed.
A
comparison
of
all
sets
of
3
or
more
data
points
with
the
Mann-Whit-
ney
test
showed
all
of
the
non-cotton
tampons
tested
affected
TSST-1
production
compared
with
both
the
still
broth
control
(P
<
0.01)
and
the
all-cotton
tampons
(P
<
0.01).
To
ensure
that
the lack
of
TSST-1
was
not
strain
specific,
an
equal
number
of
Terra
Femme
all-cotton
tampons
were
tested
using
a
second
strain
of
TSST-1
producing
S.
aureus
(No.
3984).
As
with
the
MN8
strain,
TSST-1
was
not
detected
in
any
of
the
experiments.
The
Today
contraceptive
sponge,
when
washed,
produced
1-4
Ixg/ml
of
TSST-1,
while
the
un-
washed
product
with
nonoxynol-9
produced
0-0.5
bg/ml
of
toxin.
The
nonoxynol-9
is
inhibitory
to
S.
aureus
as
evidenced
by
the
2-log
decrease
in
the
viable
bacterial
counts
of
unwashed
sponges
com-
pared
with
the
washed
product
as
well
as
with
other
products
tested.
This
finding
is
in
agreement
with
our
previously
reported
observations.
2’3
The
poly-
ester
and
carboxymethylcellulose
tampon
(Rely)
yielded
20-80
Ixg/ml
of
TSST-
when
unwashed
but
only
5-10
Ixg/ml
after
washing.
This
is
less
than
previously
reported.
Since
this
tampon
was
manufactured
almost
15
years
ago,
it
has
likely
undergone
deterioration
as
was
apparent
during
the
experiments
in
which
the
washed
product
showed
signs
of
polyester
disintegration,
discoloration,
crumpling,
and
loss
of
sponginess.
Further,
as
is
characteristic
of
the
carboxymethylcellulose,
the
Rely
tampon
could
not
be
squeezed
free
of
fluid
and
remained
in
a
gelled
state,
i.e.,
did
not
display
negative
absorption.
The
Tassaway
vaginal
menstrual
cups
are
made
from
a
non-absorbent
elastomeric
polymer.
S.
au-
reus
MN
8
produced
no
TSST-1
when
grown
in
the
presence
of
Tassaway,
whether
washed
or
un-
INFECTIOUS
DISEASES
IN
OBSTETRICS
AND
GYNECOLOGY
141
TAMPONS,
BARRIER
CONTRACEPTIVES,
AND
TSST-1
TIERNO
AND
HANNA
TABLE
I.
Chemical
composition
of
products
tested
Composition
Product
Pledget
Outer
wrap
Cord
S-L-F
Absorbency
(g)
Tampax
Regular-Compak
Rayon
Rayon
Cotton
+
6-9
Super
Cotton/rayon
Rayon
Cotton
+
9-12
Super-Plus
Rayon Rayon
Cotton
+
12-15
OB
Regular
Rayon/cotton
PNS
Rayon/cotton
+
6-9
Super
Rayon/cotton
PNS
Rayon/cotton
+
9-12
Super-Plus
Rayon/cotton
PNS
Rayon/cotton
-t-
12-15
Playtex
Reg/do/po
Rayon
None
Cotton
+
6-9
Reg/po
Rayon
None
Cotton
+
6-9
Regular-Ultimate
Rayon/cotton
Rayon
Cotton
+
6-9
Super-Ultimate
Rayon/cotton
Rayon
Cotton
+
9-12
Kotex
Regular
Cotton/rayon
PNS
Rayon/polyester/cotton
+
6-9
Super
Cotton/rayon
PNS
Rayon/polyester/cotton
+
9-12
Super-Plus
Cotton/rayon
PNS
Rayon/polyester/cotton
+
12-15
Terra
Femme
Regular
Cotton
None
Cotton
10
Super
Cotton
None
Cotton
13
Natracare
Regular
Cotton
None
Cotton
10
Super
Cotton
None
Cotton
13
CO-OP
Regular
Rayon/cotton
None
Cotton
-t-
10
Super
Rayon/cotton
None
Cotton
+
13
Rely
Super
Polyester/cmc
Polyester
Polyester
+
19
Today
Polyurethane
None
Nylon
+
20
Tassaway
Elastomeric
polymer
None
None
0
Ortho
All-Flex
Latex
None
None
0
aS-L-F
surfactant-leachable-fragrance
present;
PNS
present
not
specified;
Reg/do/po
regular
deodorant
portable;
Reg/po
regular
portable;
cmc
carboxymethylcellulose.
washed.
Ortho
diaphragms,
made
from
latex
rub-
ber,
produced
0.5
Ixg/ml
of
TSST-1
when
un-
washed
and
0.7
5
Ixg/ml
when
washed.
An
adherent
film
of
staphylococcal
growth
was
evident
on
the
surface
of
all
test
diaphragms.
TSST-1
was
de-
tected
in
the
shaker
control
broth
(3-4
Ixg/ml)
but
not
in
the
still
broth
control.
Shaker
broth
controls
produced
the
highest
viable
staphylococcal
counts
and
were
at
least
log
higher
than
the
counts
of
any
tampon
experiments.
The
viable
bacterial
counts
of
the
washed
tampons
were
slightly
higher
than
the
counts
of
the
unwashed
product.
DISCUSSION
Previous
reports
have
shown
that
vaginal
tampons,
especially
those
rich
in
synthetic
fibers,
provide
the
physical
and
chemical
parameters
that
permit
TSST-1
production
to
varying
degrees
dependent
upon
product
composition.
2-8
Numerous
physical,
chemical,
and
ecologic
factors
are
reported
to
affect
TSST-1
production
by
strains
of
S.
aureus
grown
in
the
presence
of
vaginal
products.
2-6
Currently
marketed
tampons
manufactured
in
the
United
States
(Table
3)
are
made
from
rayon
or
rayon/
cotton
blends
and
have
been
shown
to
amplify
TSST-1
in
vitro.
Since
an
all-cotton
tampon
was
not
previously
available,
comparisons
were
not
pos-
sible
until
now.
We
have
found
that
all-cotton
tam-
pons,
whether
washed
or
unwashed,
did
not
pro-
duce
detectable
TSST-1
compared
with
the
other
tampons,
which
confirms
previous
results
using
tampon-sized
pledgets
of
surgical
cotton.
2’3
This
difference
in
toxin
production
is
likely
because
cot-
ton
provides
fewer
physical-chemical
factors
that
favor
TSST-
production.
Cotton
is
less
absorbent,
has
less
surface
area,
does
not
effectively
concen-
142
INFECTIOUS
DISEASES
IN
OBSTETRICS
AND
GYNECOLOGY
TAMPONS,
BARRIER
CONTRACEPTIVES,
AND
TSST-1
TIERNO
AND
HANNA
TABLE
2.
Effect
of
products
on
the
production
of
TSST-I
Product
No.
cfu/10
-8
Unwashed
TSST-I
(lg/ml)
Range
Average
No.
cfu/lO
-8
Washed
TSST-I
(lg/ml)
Range
Average
Tampax
Regular-Compak
3
4.3
0.5-2
1.2
3
4.5
Super
2
5.8
I-4
2.5
2
6.
Super-Plus
3
3.4
2-4
3.3
3
6.0
OB
Regular
3
4.7
2-4
3.2
3
4.4
Super
3
4.2
4
4.0
3
6.
Super-Plus
3
5.4
4-8
6.7
3
7.
Playtex
Reg/do/po
3
2.2
I-2
1.3
3
4.
Reg/po
3
6.3
I-4
2.3
3
4.6
Regular-Ultimate
3
4.0
I-2
1.7
3
4.0
Super-U
Itimate
3
3.9
2-4
3.3
3
4.8
Kotex
Regular
3
3.6
I-2
1.7
2
4.2
Super
3
4.4
2-4
3.3
3
4.8
Super-Plus
3
5.6
2-4
3.3
3
6.4
Terra
Femme
Regular
20
3.
0 0
3
4.
Super
20
2.7
0
0
3
7.0
Natracare
Regular
5
2.9
0
0
3
5.2
Super
5
2.3
0
0
3
6.2
CO-OP
Regular
5
2.5
I-4
2
3
5.9
Super
5
3-4
I-2
1.4
3
6.6
Rely
Super
8
4.2
20-80
35
2
6.9
Today
3
0.03
0-0.5
<0.5
3
I.I
Tassaway
3
4.2
0
0
3
4.
Ortho
All-Flex
2
6.8
0.5
0.5
2
7.9
Broth
Controls
Shaker
(200
rpm)
15
65.0
2-4
3.0
6
68.3
Still
17
2.6
0
0
6
3.7
4
4.0
4-8
6.0
8
8.0
4-16
9.3
8-16
13.3
8-32
18.7
4-8
6.7
8
8
4-8
6.7
4-16
9.3
4-8
6.7
8 8
8-16
13.3
0 0
0 0
0
0
0
0
2-4
3.3
4-8
6.7
5-10
7.5
I-4
3.0
0
0
0.75 0.75
2-8
4.0
0
0
aReg/do/po
regular
deodorant
portable;
Reg/po
regular
portable.
trate
protein
in
aqueous
solutions,
and
provides
a
less
viscous
environment
compared
with
the
syn-
thetics
previously
removed
from
tampons
(car-
boxymethylcellulose,
polyester,
polyacrylate
rayon),
even
those
which
are
still
used
in
tampon
manufac-
ture
(viscose
rayon).
None
of
the
all-cotton
tam-
pons
tested
contain
surfactants
or
finishing
agents,
some
of
which
are
known
to
affect
TSST-1
produc-
tion.
2-4,9,10
In
the
vaginal
milieu,
many
factors
may
influ-
ence
the
polymicrobial
environment
and
the
prod-
ucts
of
bacterial
metabolism.
Independent
of
chem-
ical
composition,
factors
such
as
tampon
structure,
absorbency
or
fluid
capacity,
air
content,
and
sur-
face
area
may
affect
TSST-1
production.
3
None-
theless,
it
has
been
reported
that
cotton
actually
TABLE
3.
Marketing/distributing
country
of
vaginal
products
tested
Product
Company
Country
Tampax
Tambrands,
Inc.
United
States
OB
Personal
Products
Co.
United
States
Playtex
Playtex
Family
Products
Corp.
United
States
Kotex
Kimberly-Clark
Corp.
United
States
Terra
Femme
Bio
Business
International
Corp.
Canada
Natracare
Bodywise,
Ltd.
England/Sweden
CO-OP
CWS
England
Rely
Proctor-Gamble
United
States
Today
Whitehall
Laboratories
United
States
Tassaway
Personal
Care
Products
United
States
Ortho
All-Flex
Ortho
United
States
aNo
longer
on
the
market.
INFECTIOUS
DISEASES
IN
OBSTETRICS
AND
GYNECOLOGY
143
TAMPONS,
BARRIER
CONTRACEPTIVES,
AND
TSST-1
TIERNO
AND
HANNA
adsorbs
TSST-
1.
Cotton
loses
that
adsorbing
abil-
ity,
however,
when
combined
with
rayon.
l
It
is
likely,
therefore,
that
all-cotton
tampons
lower
the
risk
for
TSS
since
they
do
not
amplify
TSST-1
in
vitro
and
have
useful
adsorbing
affinity
for
that
toxin.
This
is
especially
important
since
menstrual
TSS
associated
with
synthetic-fiber
tampons
con-
tinues
to
occur
despite
the
decreased
absorbency;
the
elimination
of
carboxymethylcellulose,
poly-
acrylate
rayon,
and
polyester;
and
the
increased
awareness
of
the
role
of
tampons
in
TSS.
The
viable
bacterial
counts
for
washed
tampons
were
higher
than
those
of
the
unwashed
products
probably
because
inhibitory
leachables
were
re-
moved
by
the
washing
process.
Likewise,
TSST-1
increased
after
the
tampons,
except
the
all-cotton
variety,
were
washed.
When
tampons
are
placed
in
an
aqueous
solution,
they
have
been
shown
to
leach
out
various
inorganic
and
organic
chemicals
which
increases
the
viscosity
of
that
solution.
12
Washing
the
vaginal
products
prior
to
testing
more
accu-
rately
reflects
their
conditions
of
use.
Since
the
vaginal
mucosa
is
known
to
readily
absorb
chemi-
cals,
such
leachables
as
surfactants
and
other
fin-
ishing
agents
used
in
products
may
likewise
be
absorbed,
diluted,
or
inactivated
as
they
mix
with
vaginal
fluids.
2’
Since
some
leachable
chemicals
may
inhibit
toxin
production
or
the
growth
of
S.
aureus,
that
effect
may
be
mollified
by
the
washing,
hence
producing
the
slightly
higher
viable
cell
counts
and
the
significant
increase
in
TSST-1
lev-
els.
The
unwashed
product,
therefore,
does
not
reflect
a
product’s
true
propensity
for
amplifying
TSST-1
since
interference
from
some
leachable
chemicals
could
be
expressed
within
a
closed-test
system.
It
is
not
surprising
that
no
toxin
was
found
with
the
Tassaway
menstrual
cups.
They
are
made
from
an
elastomeric
polymer,
a
non-absorbent
product
that
is
apparently
inert.
In
addition,
they
do
not
allow
for
adherence
of
S.
aureus
to
their
surface,
as
occurred
with
the
latex
diaphragm.
Latex
dia-
phragms,
although
non-absorbent,
allow
for
ad-
herence
and,
as
such,