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Infusion
Fluids
as Culture
Media
E. P. CRICHTON, M.B., CH.B., F.R.C.P.(C)
Department of Bacteriology, St. Paul's Hospital, Vancouver 1, B.C.
ABSTRACT
Crichton, E. P.: Infusion fluids as culture media. Am. J. Clin. Pathol. 59:
199-202, 1973. Septicemia is a possible complication of the administration of
intravenous infusions. T h e infection may be derived from the skin or from
contaminated infusion fluid. T h e risk to the patient from contaminated infusion fluid depends on the number of organisms present and, therefore, on
the nutrient properties of the fluid. A number of commonly used infusion
fluids were tested for their growth-promoting abilities for a variety of bacteria. Some fluids did not support the growth or survival of any of the test
organisms, whereas other fluids offered selective advantages to different organisms and permitted growth at different rates and even at a pH as low as 3.5.
T H E DANGER OF INFECTION as a possible
complication of intravenous therapy is well
recognized. T h e emphasis in retrospective
and prospective investigations has been
placed on skin care, catheter care, and duration of catheterization. Relatively low infection rates have been attributed to adherence to a strict set of regulations for
skin preparation and catheter and skin
maintenance. 1 - 8
Clearly, however, infection can also arise
from contaminated infusions. T h e work
usually referred to is that of Michaels and
Ruebner, 4 who in 1953 reported two cases
of pyrexial reaction caused by bacterial
growth in the solution during infusion.
They also demonstrated that their two isolates of a coliform bacillus could grow in
physiologic saline solution and in Hartraann's solution with 5% dextrose under
conditions simulating an infusion, but not
in physiologic saline with 5 % dextrose or
in 5% dextrose alone. They attributed this
failure to grow in the latter solutions to
Received March 17, 1972; accepted for publication April 20, 1972.
the low p H of the solutions, which were
4.3 and 4.1, respectively. More recently,
several cases of fatal septicemia were traced
to contamination of the intravenous infusions at the manufacturer's plant. In one
hospital, 2 35% of the infusion sets were
found to be contaminated during use and
7 1 % of these were judged to be in unsatisfactory working condition, thus indicating
preventable circumstances. But even with
great care the risk of contamination of the
intravenous set or infusion fluid at the
time of changing the bottle or adding medication is very real. T h e degree of danger
to the patient will depend on host factors
and the nutrient properties of the fluid. As
scientific data describing these nutrient
properties are not readily available, a small
study was undertaken in an attempt to define more accurately the type of solution
which might serve as a culture medium
and lead to massive injection of organisms.
A knowledge of these growth-promoting
properties would also serve as a guide to
the frequency with which intravenous bottles and tubing should be changed.
199
200
CRICHTON
Method
A variety of organisms was selected for
the test. They were: Klebsiella, Pseudomonas aeruginosa, Serratia, Staphylococcus
aureus, Staphylococcus albus, and a Pseudomonas cepacia isolated from an infusion
bottle and blood of a baby with septicemia. The organisms were grown on a
Mueller-Hinton plate. The colonies were
touched on the surface and suspended in
distilled water. This mixed inoculum was
standardized to the Kirby-Bauer opacity
standard and diluted 1:10 in distilled
water, since distilled water is the purest
practical vehicle. A sample of this inoculum (0.05 ml.) was injected through the
rubber stopper into a standard intravenous
infusion bottle (Cutter Laboratories, Inc.,
Berkeley, Calif. 94710, USA). After 15
min., fluid was withdrawn from the bottles using a different Old Tuberculin syringe for each bottle, and two drops were
seeded onto a Mueller-Hinton plate. The
bottles were left at room temperature to
simulate "in use" conditions and cultured
again at 7 hr., 24 hr., 48 hr., and 144 hr.
T h e plates were incubated overnight and
the numbers of colonies counted and recorded. The types of organisms that multiplied were identified by colonial morphology. When an infusion showed an overgrowth of Gram-negative bacilli, an aliquot
was inoculated onto a mannite salt agar
in order to detect the presence of staphylococci. Additionally, Staphylococcus aureus
and 5. albus were inoculated into the fluids
alone and together to determine whether
these cocci could multiply in the absence
of interference by Gram-negative bacilli.
The pH of the solution was measured
with a glass electrode before and immediately after inoculation. The test series was
done on three different occasions.
Results
The findings in the diree tests were similar. Only minor discrepancies occurred and
A.J.C.P.—Vol.
59
the less favorable findings are recorded in
Table 1. Significant increases in bacterial
numbers occurred at different time intervals
with different solutions. Also, the solutions
provided selective advantages to different
organisms, and the organisms which were
recognizable morphologically are recorded.
In most cases in which growth occurred,
Serratia was present, but in 10% invert
sugar in water the Pseudomonas cepacia
multiplied much better. Staphylococci, however, could not be isolated from any of the
fluids with a growth of Gram-negative bacilli. Staphylococci did not multiply when
they were inoculated into the fluids alone,
and from most of these fluids the two-drop
culture was negative after 72 hr. The pH
of the solution was not changed by the addition of one drop of suspension, and some
of the bacilli grew well in solutions of
low pH, as indicated.
Discussion
The testing system attempts to simulate
"in use" conditions, but by necessity the
inoculum had to be larger. The organisms
were selected as representatives of likely
contaminants and isolates from patients
with bacteremia or septicemia. It is interesting that the Staphylococci did not multiply in any of the fluids. This would explain the repeated observation that Gramnegative bacilli are isolated from contaminated infusions more frequently than
Gram-positive organisms; the original contamination may have included Gram-positive organisms, but they did not survive. It
is also tempting to speculate that, in the
absence of other host infection, colonization of the catheter tip by Gram-positive
cocci arises from skin organisms or contamination at the skin site, whereas Gramnegative bacilli may originate in the infusion fluid.
Noteworthy findings are the different
rates of multiplication of different organisms in different solutions, indicating the
February 1973
INFUSION FLUIDS AS CULTURE MEDIA
201
Table 1. Nutrient Properties of Intravenous Solutions
G•owth afterf
OverpH growth*
Distilled water
NaCl inj.
Lactated Ringer's
M/6 Na Lactate inj.
Z\% Dextrose + 0.3% NaCl
3J% Dextrose + 0.3% NaCl + KC1
5% Dextrose + 5% alcohol
5% Dextrose electrolyte 48
5% Dextrose in saline
5% Dextrose in H20
5% Dextrose in lactated Ringer's inj.
5% Dextrose + 5% protein hydr. in H20
10% Dextrose in saline
10% Dextrose in H20
10% Dextrose + 6 mEq. KC1 + 8 mEq.
NaHCOa
20% Dextrose U.S.P.
50% Dextrose in H20
10% Invert sugar in H20
10% Invert sugar in NaCl
10% Invert sugar in electrolyte 2
10% Invert sugar + 0.3% KC1
+ 0.45% NaCl
5% NaHCOs in H20
Polysal (bal. replacement electrolyte)
Polysal + 5% dextrose
Rheomacrodex 10/W/V in N saline
Rheomacrodex 10/W/V in dextrose
Amigen 5% in 12.5% levugen,
2.4% alcohol
Intralipid 10%
Polyonic M 900
15 min. 7
hr.
24
hr.
48
hr.
72
hr.
144
hr.
CD
CD
CD
CD
CD
CD
—
4
CD
CD
4
—
18
21
17
11
7
1
6
18
11
16
17
4
9
20
11
13
17
9
14 C D
6 46
4 37
4 CD
1
2
4
14
6
1
16 C D
14
8
11
6
8
6
16 10
C
C
C
C
C
C
D
D
D
D
D
D
—
9
2
CD
15
6
4
CD
CD
CD
CD
CD
CD
CD
—
3
CD
CD
8
6
CD
CD
8.0 S, K, Ps
3.7
3.4
3.5 Pb, S
3.2
4.5
40
16
17
26
21
5
28 C D
5
3
1
5
3
8
2
16
5
8
CD
—
5
6
—
5
CD
—
4
CD
—
3
—
—
CD
—
6
3.3
7.9
5.8
5.4
4.2 S, K
4.1 S, K
16
11
7
12
22
14
5
1
10
10
15
5
7
8
16 C D
60
21
2
10
5
13
CD
CD
—
10
8
13
CD
CD
—
3
1
9
CD
CD
5.1
6.5 S, K, Pb
3.8
2
9
17
3
1
9 CD
I
1
—
CD
—
—
CD
—
—
CD
—
7.0
6.1
6.6
6.8
4.8
4.8
4.0
4.5
4.4
4.5
4.6
5.1
4.1
4.4
Pb,S
S, K
S,Ps
S
S
s
S, K
s
s
s
CD.
* K = Klebsiella, S = Serratia, Ps = Pseudomonas aeruginosa, Pb = a Pseudomonas cepacia from a baby with septicemi
t The arable numerals indicate the number of colonies cultured per plate. The + symbols indicate increase in growth beyond
a countable number (approximately 100) and the number of positives the relative amount of increase, with (j^jj) representing
confluent growth.
202
CRICHTON
need to test a variety of organisms, and
the growth of Serratia and the Pseudomonas cepacia in a pH as low as 3.5 and
in commercial distilled water.
The findings apply only to the solutions
tested without any further additions, and
only to the organisms tested. Contamination problems and prevailing organisms
vary from area to area and time to time,
and no test system of this nature could be
comprehensive.
The practical conclusions apply to the
need for changing not only the bottle but
the tubing. The tubing is important, as it
has been shown that multiplication of organisms can occur even if the bottles are
changed to sterile solutions.4 If multiplication of bacteria did not occur in the fluid,
then there would be no need to change the
tubing. If, on the other hand, the infusion
A.J.CP.—Vol.
59
fluid is a good nutrient, then the decision to change the administration set would
depend on the rate of increase of organisms
and the likelihood of contamination of the
fluid, which again would depend on the
care with which the set was handled and
the number of times the circuit had been
broken or medication had been injected.
Acknowledgment. Miss E. Underhill provided
technical assistance.
References
1. Corso JA: Maintenance of venous polyethylene
catheters to reduce risk of infection. JAMA
210:2075-2077, 1969
2. Duman RJ, Warner JF, Dalton HP: Septicaemia
from intravenous infusions. N Engl J Med 284:
257-260, 1971
3. Fuchs PC: Indwelling intravenous polyethylene
catheters. JAMA 216:1447-1450, 1971
4. Michaels L, Ruebner B: Growth of bacteria in
intravenous infusion fluids. Lancet 1:773-774,
1953