Transfusion Medicine, 2009, 19, 0–0
ORIGINAL ARTICLE
Blood surveillance and detection on platelet bacterial
contamination associated with septic events
J. C. Hsueh,* C. F. Ho,* S. H. Chang,* F. Z. Pan,* S. C. Chen,* M. D. Shi† & S. T. Chien*‡
*Department of Pathology, Kaohsiung Armed Forces General Hospital, Kaohsiung, Taiwan, †Department of Pathology and Laboratory Medicine,
Yongkang Veterans Hospital, Taiwan, ‡Department of Medical Technology, Fooyin university, Taiwan
Received 6 October 2008; accepted for publication 30 May 2009
SUMMARY. The objective of this study was to evalu-
ate the risk of transfusion-associated septic events in
Taiwan. In Taiwan, most blood components are pro-
vided from blood centres; so platelet (PLT) bacterial
contaminations are rarely reported. The study’s aim is
to investigate the prevalence of PLT bacterial contami-
nation in Kaoshiung Armed Forces General Hospital by
using BacT/ALERT system for routine screening.
A total of 82 apheresis and 2256 whole
blood-derived PLT units were tested. A measured quan-
tity of 1 mL aliquots were taken as samplings from all
blood bag tubing of PLT units, and then further incubated
in a bacterial detection system (BacT/ALERT). The
subcultures of true-positive bottles underwent bacterial
identification by using Vitek system, microscopic obser-
vation and culture-based methods. Eight units (0·34%,
8 of 2338) were found to have bacterial contamina-
tion. The true-positive rate of the whole blood-derived
Platelet (PLT) bacterial contamination is an ongoing
and critical problem, and it is thought to be highly
associated with transfusion morbidity and mortality
worldwide. Therefore, the American Association of
Blood Banks (AABB) adopted a new standard in 2004,
which requires transfusion service and blood banks to
have routine screening procedures in order to reduce
incidents of bacterial contamination of PLTs. Preserv-
ing PLT at room temperature may inversely create an
opportunity for initial bacterial inoculation growth in
Correspondence: S. T. Chien, Department of Pathology, Kaohsiung
Armed Forces General Hospital, No. 2 Chung Cheng 1st Road,
Kaohsiung City, Taiwan.
Tel.: +886-7-7496751; fax: +886-7-7491013
e-mail: chienstkl@gmail.com
© 2009 The Authors
Journal compilation © 2009 British Blood Transfusion Society
doi: 10.1111/j.1365-3148.2009.00949.
and apheresis PLTs was 0·31% (7 of 2256) and 1·22%
(1 of 82), respectively. Six microorganisms were iden-
tified with the most dominate being Staphylococcus
epidermidis. One case of transfusion-associated sepsis
was confirmed; in addition, the holding period of PLTs
(F = 4·522, P = 0·034) and positive detection of PLT
bacterial contamination (F = 46·605, P < 0·001) were
associated with post-transfusion sepsis. Thus, in this
study, although the transfusion-associated septic event
was rarely found and PLT units were provided from
blood centres, bacterial screening was necessary to safely
quarantine the transfusion. The holding period of PLT
units should be no more than 4 days in order to avoid
possible bacterial contamination.
Key words: bacterial contamination, blood surveillance,
platelet, transfusion.
great amounts. Therefore, several reports have been
documented that state PLT units must be used within
a 5-day storage period (Braine et al., 1986; Blajch-
man et al., 2004). The older PLT units may be con-
sidered to have higher bacterial inoculation and thus
result in sepsis. Nevertheless, the adequate period of
storing PLT is between 5 and 7 days in the United
States, and conditionally, prolonged to 7 days when
culture-based method is applied for qualification. In
contrast, culture-based method often needs a conven-
tional period to determine PLT bacterial contamina-
tion. Therefore, PLT bacterial contamination is still
estimated at 0·35% when routine culture-screen was
implicated for the PLT units that had been stored for
5–7 days (Lee et al., 2003). Based on these findings,
the shelf-life of PLT units was suggested to be 5 days
1
2 J. C. Hsueh et al.
in order to avoid bacterial contamination. The posi-
tive rate of PLT bacterial screening was reported to be
around 1/2000 to 1/3000, which was estimated by using
different approaches to survey the prevalence of bacte-
rial contamination in single-donor PLT units (apheresis)
and random-donor PLT units (whole blood-derived)
(Kleinman et al., 2006; Yomtovian et al., 2006). Basi-
cally, the ratio of PLT consumption between aphere-
sis units vs. whole blood-derived PLT units (56·7 vs.
43·3%) was different from that of the United States.
This discrepancy might be due to cultural difference
and medical system in blood donation. According to
the previous report, it was suggested that bacterial con-
tamination in PLTs occurred around 1/1600 to 1/4500
in Taiwan (Kaohsiung Blood Donation Center of Tai-
wan Blood Services Foundation, 2001–2003). Despite
the difference in the bacterial detecting systems that
were utilized for PLTs’ surveillance, the PLT bacterial
contamination has not decreased worldwide. There is
an account for approximately 17% of severe episodes
in transfusion contamination. Moreover, the ratio of
transfusion-associated septic events to PLT transfusion
was estimated at 1:25 000 (1:13 000–1:100 000) in
United States before 2004 (Kuehnert et al., 2001; Ness
et al., 2001; Perez et al., 2001).
Actually, patients who received contaminated PLTs
might appear with symptoms such as chills, rigors or
fever, where several clinical indications could easily be
ignored and become irrelevant to bacterial contamina-
tion. First of all, it might bring complexity to distin-
guish aforementioned symptoms which were caused by
transfusion or adverse effects from antibiotic therapy,
when patients exhibited neutropaenia or fever. Sec-
ondly, a positive result from the culture-based method
is insufficient to support a true case of transfusion-
associated sepsis. Therefore, an erroneous hypothe-
sism, such as PLT bacterial contamination, never prac-
tically evoked clinical outcomes, might easily to be
assumed due to above reasons. The finding of clin-
ical evidence was necessary to verify true-positive
cases of transfusion-associated septic event. Several
common PLT contaminated bacteria have been iden-
tified, which include coagulase-negative Staphylococci
(CNS), Bacillus spp. (Kleinman et al., 2006), Staphy-
lococcus spp. (Fang et al., 2005), Salmonella species,
Escherichia coli, Leclercia adecarboxylata (Davenport
& Land, 2007) and Serratia marcescens (Lin et al.,
2004). Moreover, Yersinia enterocolitica was docu-
mented as a common, isolated, transfusion-associated
bacterium in United States. Nevertheless, there have
only been a few research articles that address the
issues of the transfusion-associated sepsis event which
is highly relevant to PLT bacterial contamination. In
this study, we conducted blood surveillance of PLTs
Journal compilation © 2009 British Blood Transfusion Society, Transfusion Medicine, 19, 0–0
to evaluate the prevalence of bacterial contamination
and transfusion-associated sepsis events. In addition,
we examined whether the holding period of PLTs,
PLT types (apheresis vs. whole blood-derived), positive
detection of PLT bacterial contamination and num-
bers of transfused blood bag had any relation to the
transfusion-associated sepsis.
MATERIALS AND METHODS
The source of PLT units
All blood components were provided from the local
blood centre, the Foundation of Blood Donation Ser-
vice in Taiwan. According to the monthly bulletins
from the local blood centre, it was reported that random
collected PLTs (∼ 7·5%) were subjected to bacterial
detection system (BacT/ALERT, bioMérieux, Durham,
NC, USA) for bacterial screening, whereas other PLT
units were released to hospitals for further usage. Our
hospital received a total of 2338 PLT units, includ-
ing 82 apheresis and 2256 whole blood-derived units
between February 2006 and June 2007. The averaged
holding period for PLT unit was 2·5 days (data not
shown). There were 4 apheresis and 485 whole blood-
derived units of PLTs having a holding period of 1 day;
36 units of apheresis and 807 units of whole blood-
derived PLTs of 2 days; 20 units apheresis and 521
units of whole blood-derived PLTs of 3 days; 11 units
apheresis and 311 units of whole blood-derived PLT
of 4 days; and 11 units apheresis and 145 units whole
blood-derived PLTs of up to 5 days in storage.
Detection and identification of PLT bacterial
contamination
To ensure PLT transfusion safety and quality control,
each PLT unit was subjected to bacterial screening
by which 1 mL aliquots from blood bag tubing were
sampled. First, the collected blood bag tubing was
cleaned with providone iodine and 70% alcohol in lam-
inar flow, and then 1 mL aliquots of all PLT units
were immediately sampled, inoculated into aerobic
and BacT/ALERT anaerobic culturing bottles. These
BacT/ALERT bottles were continuously monitoring for
5–6 days, or till positive results were indicated. The
suspected positive bottles were subjected to culturing
tests by using blood agar plate/eosin methylene blue
biplate agar (BAP/EMB) (Becton, Sparks, Maryland,
USA), chocolate agar plate, CDC BAP (CDC anaer-
obe 5% sheep blood agar plate), which were cultured
for 18–24 h (37◦ C±1, 5% CO2 ). Furthermore, they
were prepared on smear specimens on Gram stain-
ing. The positive cultures were directly observed with
© 2009 The Authors

a microscope and specified from their characteristics
through morphology. The verified-positive subcultures
of both culturing tests and microscopic observation
were additionally substantiated by using the vitek sys-
tem (bioMérieux, Durham, NC, USA). The turbidity
of each bacterial suspension was adjusted to match
a McFarland (0·5) standard in a sterile solution of
sodium chloride (0·45%) within 30 min to avoid vari-
ation in thickness. Then vitek GPI was applied to per-
form samples in filling, sealing and incubating till the
data were generated. The true case of PLT transfusion-
associated sepsis was defined as successfully detecting
identical bacteria with the same resistant pattern of
antimicrobial susceptibility test by using different sam-
ple sources including the subcultures of transfused PLT
units and recipient’s venous blood. In addition, the
recipients were observed for 5–7 days to verify that
their clinical outcomes did correlate with PLT transfu-
sion sepsis.
Susceptibility testing
The bacterial suspension was transferred in 200 μL of
1·8-mL sterile saline (0·45%), and then immediately
inoculated into GPS-110 cards for bacterial identifi-
cation by using the Vitek system. There were 8 iso-
lated tests against 17 antimicrobial agents including
amikin, ampicillin, sulbactam/ampicillin, ceftazidime,
cefepime, ceftriaxone, cefuroxime, ciprofloxacin, gen-
tamycin, imipenem, piperacillin-tazo-bactam, SXT, lev-
ofloxacin, cefotetan, cefuroxime-Na, ceftazidime and
piperacillin. Three control strains were used as refer-
ence and were inoculated in each run to ensure the
adequacy and antibiotic ranges. The MICs were gen-
erally classified as susceptible (S), intermediate (I) or
resistant (R), according to the breakpoints which were
recommended by the NCCLS (1999).
Statistical analysis
The anova test was applied to distinguish the differ-
ences between two PLT types. The holding periods of
PLTs and PLT types were analyzed by using univariate
linear regression of general linear model in spss ver-
sion 15·0 for Windows software (SPSS, Chicago, IL,
USA) to determine whether they were associated with
PLT bacterial contamination. In addition to the hold-
ing period of PLTs and PLT types, positive detection
of PLT bacterial contamination and numbers of trans-
fused blood bag in relation to the transfusion-associated
sepsis were also determined in the same way. The com-
parison of proportions was performed with two-sided
Fisher’s exact test to elucidate whether the difference in
positive detection of PLT bacterial contamination was
© 2009 The Authors
Journal compilation © 2009 British Blood Transfusion Society, Transfusion Medicine, 19, 0–0
Bacterial screening of PLT contamination 3
significant between the two subgroups of PLT incubat-
ing duration (1–3 days vs. ≥4 days). The odds ratio
of the dichotomous subgroup to PLT bacterial con-
tamination was estimated. Differences were considered
statistically significant when P value was less than
0·05.
RESULTS
In total, 2338 PLTs comprising of 2256 units of
whole blood and 82 apheresis units were received
and tested to control bacterial contamination. The PLT
units were transfused into 323 patients who were given
follow-ups in a short period. The data for 12 months
of bacterial screening are presented in Table 1. The
true-positive rate of the whole blood-derived PLTs
and apheresis units had an occurrence rate of 0·31%
(7/2256) and 1·22% (1/82), respectively. The eight
suspected positive cases were all true-positive with
corresponding bacterial isolates which were completely
(100%) identifiable from Gram staining and Vitek
system. The results of the bacterial identification and
the pattern of antimicrobial susceptibility test are
shown in Table 2. All eight isolates were associated
with skin or endogenous origins, which were identified
as Sta. epidermidis, Sta. warneri, Sta. haemolyticus,
Sta. hominis, Gram-positive bacilli (GPB) and Gram-
positive cocci (GPC). The difference between the two
types of PLTs was compared by using anova test.
Accordingly, a significant difference was observed in
the numbers of blood bag (F = 80·285, P < 0·001,
data not shown), but there were no evident difference
found in the holding period of PLTs, culturing days of
subculture of positive bottle and positive detection of
PLT bacterial contamination. It is plausibly that more
blood bags should be implicated for transfusion when
whole blood-derived PLTs are used. This difference in
numbers of blood bag between two different PLTs may
simply be due to the averaged PLT units in each bag
of the whole blood-derived PLTs being less than that
of apheresis PLTs.
Table 1. The positive rate of PLT bacterial contamination
screening
PLT type True-positive rate
Whole blood-derived 7/2256 0·31%
Apheresis 1/82 1·22%
Overall 8/2338 0·34%
The result of bacterial screening of PLTs was shown as above, and
only the true-positive rate was indicated due to the contaminated
bacteria that were successfully isolated from the subcultures of eight
BacT/ALERT positive cases.
 4 J. C. Hsueh et al.

Table 2. The general characteristics of the isolated PLT contaminated bacteria in this study

AST pattern

PLT con- Duration of Numbers of

Culturing taminated Blood antibiotic transfused
Case days bacteria culture Antibiotics therapy blood bag SAM CZ CC E GM OX P RA TE SXT VA LVX MXF

1 3 Sta. epi Sta. epi cefa 2 days 1 R R R R R R R S R S S S I

2 2 Sta. warneri Augmentin 1 day 7 R R S S S R R S S S S S S
3 2 Sta. hae Recef 2 days 12 S S S S S S R S S S S S S
4 3 GPB Cefazolin 3 days 24 S S R S S S S S S S S S S
5 7 GPB Cefa 12 days 24 S S R S S R S S S S S S S
6 5 GPC Tienam 1 day 1 S S S S S R S S S S S S S
7 5 GPB Cefazolin 2 days 9 S S R S S S S S S S S S S
8 3 Sta. hominis Gentamycin 2 days 8 S S S R S S R S R R S S S

Cefa, Cefadroxil; Recef, cephradine; Tienam, imipenem and cilastatin sodium (MSD); SAM, Ampicillin-sulbactum; CZ, cefazolin; CC, clindamycin; E, erythromycin; GM, gentamycin; OX,
oxacillin; Sta. epi, Staphylococcus epidermidis; Sta. warneri, Staphylococcus warneri ; Sta. hae, Staphylococcus haemolyticus; Sta. hominis, Staphylococcus hominis; GPB, Gram-positive bacilli;
GPC, Gram-positive cocci; AST, antimicrobial susceptibility test; S, susceptible; I, intermediate; R, resistant.
The only transfusion-associated sepsis case was identified by discovering identical bacteria isolated from both BacT/ALERT subculture and the peripheral blood of the recipient. The identifiable
microorganism was Sta. epidermidis which is indicated in boldface type. In addition, the drug-resistant pattern of antimicrobial susceptibility tests of BacT/ALERT subculture and blood culture
was consistent.

© 2009 The Authors

Journal compilation © 2009 British Blood Transfusion Society, Transfusion Medicine, 19, 0–0

Furthermore, we performed univariate analysis of
generalized linear model to ascertain whether the hold-
ing period of PLTs and PLT types was associated with
PLT bacterial contamination. It was suggested that the
holding period of PLTs might be an independent factor
rather than the PLT types that were associated with
PLT bacterial contamination (F = 9·548, P = 0·002,
data not shown). There was no statistically significant
association between PLT subtypes and bacterial con-
tamination (F = 0·515, P = 0·473, data not shown). In
addition, we also performed the analysis to determine
whether the holding period of PLTs, PLT types, positive
detection of PLT bacterial contamination and numbers
of transfused blood bag were relevant to transfusion-
associated sepsis. According to the results, the hold-
ing period of PLTs (F = 4·522, P = 0·034, data not
shown) and positive detection of PLT bacterial contam-
ination (F = 46·605, P < 0·001, data not shown) were
independently associated with transfusion-associated
sepsis. Nevertheless, it is suggested that both the num-
bers of transfused blood bag (F = 1·387, P = 0·240,
data not shown) and PLT types (F = 1·390, P =
0·239, data not shown) were not correlated to the
transfusion-associated septic events. According to the
results of two-side Fisher exact test shown in Table 3,
the shelf-life of PLTs was suggested to be within
4 days of storage. It was observed for having a sig-
nificant difference in detecting PLT bacterial contam-
ination between two groups of PLT holding period
(1–3 days vs.≥4 days, P < 0·003). The PLTs that had
a storage period longer than 4 days were estimated
for 3·09 times risk to be associated with bacterial
contamination when compared to those preserved less
than 4 days. Besides, one PLT bacterial contaminated
case was finally identified as true transfusion-associated
septic event as identical results of antimicrobial sus-
ceptibility test were obtained by detecting subcul-
tures of transfused PLTs and patient’s peripheral blood
(Table 2).
Table 3. The result of two-side Fisher’s exact test after
comparing the difference between the two subgroups of the
holding period of PLTs
Holding period of PLTs
1–3 days ≥4 days P value
Positive rate 2/222 6/71 0·003**
OR 0·30 3·09
95% CI 0·19–0·48 0·93–10·27
The denominator was the number of recipients.
∗∗
indicates P < 0.01
© 2009 The Authors
Journal compilation © 2009 British Blood Transfusion Society, Transfusion Medicine, 19, 0–0
Bacterial screening of PLT contamination 5
DISCUSSION
Our overall true-positive rate for PLT bacterial con-
tamination (0·34%, 8/2338) is much higher compared
to the report from American Red Cross (ARC; 0·017–
0·019%) (Fang et al., 2005), but the prevalence of PLT
bacterial contamination is lower than the results from
European reports (0·38–0·72%) (Munksgaard et al.,
2004; te Boekhorst et al., 2005), which were based
on prospective BacT/ALERT culturing of whole blood-
derived and apheresis PLTs. The plausible reason for
having a higher positive detection rate of PLT bacte-
rial contamination was due to our PLT units that were
received from a blood centre that used bacterial screen-
ing only randomly by culturing test. Moreover, the
higher positive detection rate in apheresis PLTs seem
more likely influenced by a small sample size (n = 82)
in comparison with that of whole blood-derived PLTs
(n = 2256) used in this study. Besides, some of aphere-
sis PLTs received from the blood centre before 2007
was not completed with random bacterial screening.
This latent factor may result in an apparently higher
positive detection rate.
The major microorganisms identified in this study
were endogenous or transient skin flora which con-
curs with the consistent result of other publications
(AuBuchon et al., 2002; Brecher et al., 2003; Fang
et al., 2005). In addition, the contaminated bacteria
were coagulase-negative and were identified as Staphy-
lococcus spp., GPB and GPC. These results were simi-
lar to those reported by ARC and other research groups.
However, we did not identify any true-positives of
Gram-negative organism’s contamination in this 1 year
investigation. This finding is different from ARC expe-
rience where Gram-negative bacteria contamination
accounted for 17·7% in true-positive isolates. Over-
all, our finding has a similarity to more recent liter-
ature, which had true-positive isolates detection rate
of 0·0074% (one in 13 579) by using whole blood-
derived PLTs and BacT/ALERT system in local hos-
pitals in the United States (Kleinman et al., 2006).
In this study, one transfusion-associated sepsis was
found based on identical antimicrobial susceptibility
pattern of the isolated bacteria achieved by detecting
the subcultures of transfused PLTs and patient’s periph-
eral blood. The contaminated bacterium was identified
as Sta. epidermidis, which is frequently isolated from
skin and mucous membrane in humans. Only one of
eight positive PLT bacterial contamination cases was
determined to be associated with transfusion-associated
sepsis. This PLT-transfused patient underwent cardiac
surgery, and Sta. epidermidis was isolated from his or
her venous blood. It is known that Sta. epidermidis is a
major pathogen that might induce endocarditis during
6 J. C. Hsueh et al.
open-heart surgery. Therefore, the carditis might not
be caused by simplistic reasons as previous described
(Munksgaard et al., 2004). Other than bacterial con-
tamination in valve replacement procedures and deep
internal wound infections, it was suggested that trans-
fusion of bacterial contaminated PLTs should be con-
sidered as a risk factor of endocarditis. Although PLT
transfusion-associated septic events were rarely dis-
covered in this study, the negative findings might be
attributed to several reasons, such as the successful-
ness of antibiotic therapy, inattentiveness of bacterial
detection and insufficient culturing duration for bac-
teria growth. According to the experiences of bacte-
rial spiking studies, a 24-h incubation was suggested
for sufficient bacterial growth, such as Sta. epider-
midis, when BacT/ALERT system with aerobic bottle
was applied (McDonald et al., 2001; Ramirez-Arcos &
Goldman 2005). Nevertheless, an extended incubation
period might be necessary, although initiative inocula-
tions are too small to adequately detect the PLT bac-
terial contamination. Theoretically, a prolonged period
of bacterial culturing might provide better compara-
tive results to identify more true-positive cases, which
could support the association between PLTs’ bacte-
rial contamination and transfusion-associated events.
According to our finding, the positive detection of
PLT bacterial contamination did have a link to the
transfusion-associated sepsis, even though the case was
rare. Unexpectedly, it had been suggested that the PLT
bacterial screening by culturing-based methods might
reduce the risk, but it could not totally eliminate sep-
sis caused by PLT bacterial contamination (Benjamin
& Mintz, 2005). Therefore, more intensive work with
a molecular approach (e.g. PCR based methodology)
might be necessary to provide better transfusion safety.
In this study, the holding period of PLTs was found
as a major determinant to PLT bacterial contamination,
as well as transfusion-associated sepsis. This finding
was partially agreed with the current opinion of PLT
storage in which the shelf-life of PLTs was suggested to
be remaining within 5 days (Lee et al., 2003). Accord-
ing to the result, we suggest a 4-day room tempera-
ture preservation of PLTs that may assure transfusion
safety. Furthermore, the positive detection of PLT bac-
terial contamination by using multiple verifying proce-
dures in our system also had an impact on identifying
transfusion-associated sepsis. In summary, we effec-
tively performed bacterial test on 82 apheresis and 2256
whole blood-derived PLTs over a 12-month period in
this study. Eight cases of bacterial contaminated PLTs
were found, and one transfusion-associated sepsis case
was verified. We suggest that bacterial contamination
screening is necessary to quarantine the transfusion
Journal compilation © 2009 British Blood Transfusion Society, Transfusion Medicine, 19, 0–0
safety in a local hospital when PLT units were pro-
vided from blood centres. We hope these findings will
improve quality control and reduce bacterial contami-
nation of blood transfusion in Taiwan.
ACKNOWLEDGMENT
This work was supported by grant No. 9632 from the
Division of Outpatient Services of Kaoshiung Armed
Forces General Hospital, Kaoshiung, Taiwan.
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