Radiometric Blood Culture Techniques
J. C. McLAUGHLIN,* P.HAMILTON,J. V. SCHOLES,t AND R. C. BARTLETT Divisionof Microbiology, Department of Pathology, Hartford Hospital, Hartford, Connecticut 06115
Received 19 May 1983/Accepted 28 July 1983
Thelysis-centrifugation technique (ISOLATOR; E. I. du Pont de Nemours & Co., Wilmington, Del.) and the radiometric blood culturetechnique (BACTEC;
Johnston Laboratories, Inc., Cockeysville, Md.) were compared on 1,000blood
cultures. Atotal of 16 ml of blood wasdistributed: 8 ml intoan ISOLATOR 7.5 microbial tube and 4 ml each into BACTEC 7C and 8B bottles. The concentrate
from the ISOLATOR tubes was inoculated underalaminar-flow hood onto two
sheepbloodagarplates (one incubated inCO2and one incubatedanaerobically), one chocolate agar plate, and one brain heart infusion agar plate. Of 91 blood
specimensobtained thatyielded clinically significant organisms, 52werepositive
by both systems, 27were positive bytheISOLATOR system only, and 12were
positive by the BACTEC system only. From the positive blood specimens, 97
clinically significant organisms were isolated: 57 by both systems, 27 by the
ISOLATOR system only, and 13 by the BACTEC system only. Of the 57
organisms detected by both systems, 28 were detected simultaneously, 13 were
detected earlierby the ISOLATOR system, and 16were detected earlierby the
BACTEC system. Isolated colonies were obtained earlier by the ISOLATOR
systemin 40casesandbythe BACTEC systemin 5cases.Organismsdetermined tobe contaminants by thoroughchart reviewwereisolated from 138ISOLATOR
tubes. In98instances,these were represented byonecolony ofStaphylococculs epidermidis, alpha-hemolytic streptococci, or diphtheroids. The ability to deter-mine CFU per milliliter with the ISOLATOR system did not help differentiate
clinically significant organismsfrom contaminants.
ISOLATOR(E. I. duPont de Nemours &
Wilmington, Del.) bloodculture system
lysis of blood, subsequent
aspiration ofthe concentrate,
appropriate media.Dorn et
Staphylococcusaureus, Pseudomonas aerugin-osa,
(L-C)method. Kiehn et al. (T. E.
Edwards,and D. Armstrong, Abstr. Annu. Meet. Am.
C82,p. 325)reported that
significantlymoreEscherichia coli and yeast isolateswererecoveredby the L-C
techniqueand that more Pseudomonas iso-lates were recovered in a conventional blood
technique. Henryet al. (N. K. Henry, C. A.
Wright,W. R. Wilson,
Thompson,and J. A. Washington, Pro-gramAbstr. Intersci. Conf. Antimicrob. Agents
tPresent address: Department of Pathology, St. Luke's-Roosevelt Hospital Center,NewYork, NY 10025.
22nd, Miami Beach, Fla.,abstr. no.
frequency of isolationof S. aureus and
Candidaspp. and a decreased time to
detection of these organismsand
etal. (M. J. Fojtasek, T. M.Abbott, J. M. Mat-sen, and M. T.
Kelly,Abstr. Annu. Meet. Am.
C129,p. 293) also
conventional two-bottlesystemwith the L-C
92%were detected by the L-C
tech-nique and only 78%were detected by the con-ventional method and that the isolated colonies
were found 24 h earlier by the L-C method. Gerlachetal. (E. H.Gerlach, R. C. Weller, and
R. J. Taylor, Abstr. Annu. Meet. Am. Soc. Microbiol. 1982,
C127,p.292), however, report-ed that, with extensive subculturing, a three-bottle system recovered
92%of 104 positive
cultures, comparedwith 76% recovered by the L-C system. Isenberg compared the L-C, the BACTEC
radiometric,and the broth culture
approacheson 996 blood samples. Significant
on February 7, 2020 by guest
organismswere isolated from 9.7% of theblood specimens by the L-Ctechniqueandfrom6.4% by the radiometric technique (2).
the yield of microorganisms from 1,000 blood specimens by the L-C technique (ISOLATOR)
and the radiometric blood culture technique (BACTEC; Johnston Laboratories, Inc., Cock-eysville, Md.). The study was also designed to comparethe time todetect andisolateorganisms bythe two systems and thecontamination rate
of each system.
Bloodcollection,inoculation, and incubation. Blood specimens includedin this study were collected only by members of thephlebotomyteamafter appropriate disinfection of the venipuncture site. Children under the ageof 6 years were notincluded. A total of 16 ml of blood was drawn: 8 ml wasinoculated into an ISOLA-TOR7.5microbialtube, and4ml eachwas inoculated into two BACTEC vials(8Bhypertonic aerobic and 7C anaerobic). BACTEC bottles were incubated at 35°C within 2 h of blood collection. Bottles were examined visually daily, and aerobic bottles were sampled on the BACTEC 460 ondays 1, 2, 3, and7, and anaerobic bottles were sampled on days 2, 4, and 7. For the purposes of this study, all BACTEC bottles were subcultured at 24h. Broth from the 8B bottles was subculturedto achocolate agarplateincubated aerobi-cally, and broth from the 7C bottles was subcultured to
achocolate agarplate incubated aerobically and to a blood agarplate incubated anaerobically. These plates were held for 48 h.
AllISOLATOR tubeswereprocessedwithin 2 h of blood collection. After a thorough mixing of blood with tubeingredients, the tube was centrifuged (3,000
x g) for 30 min. The tube was vented, and the
supernatantwasremoved anddiscarded. The tube was placedon aSuper-Mixer (Lab-Line Instruments, Inc., Melrose Park, Ill.), the yellow stopper was disinfect-ed, and the 1.5 ml of concentrate was aspirated. Approximately 0.4 ml of concentrate was used to
inoculate each of four plates: 2 sheep blood agar plates,oneincubatedfor4days aerobicallyin5%CO2
andoneincubated for6days anaerobically in Gas-Pak jars (BBLMicrobiology Systems,Cockeysville, Md.);
onechocolate agarplateincubated for4days aerobi-cally in 5% C02; and one brain heart infusion agar plate incubated for 8 days aerobically. Plates were
inoculated by the drop, tilt, and streak technique.
Plates were examineddaily todetermine the timefor isolationand the number ofCFU per milliliter.
Chart review. A thorough chart review was per-formedonpatientswhose bloodyieldedlow numbers
consid-eredcontaminants. Allbutfive charts wereavailable forreview. Information from laboratory records and
communication with physicians treating the patients provided somedata in thecasesfor whichcomplete recordswereunavailable. Specificdataobtainedfrom each chart and systematically recorded included: (i)
otherpositiveculturesofblood or othersites;(ii)the
presenceofpredisposingfactors and sources of
infec-tions; (iii)antibiotictherapy, especiallyatthe time of
culture; (iv) the presence of fever greater than 100°F (37.8°C) or chills; (v) leukocytecount elevation; (vi) type and time of surgical procedures; and (vii) evi-dence ofpneumonia, urinary tract infection, wound infection, endocarditis, abdominal sources of infec-tion, or otherinfection based on clinical and labora-torydata, including cultures.
Based on these data and on the record of other isolatesfrom the patient, eachisolate wasclassified as a contaminant, transient bacteremia, or septicemia. For purposesof thisanalysis, the term nonpathogens refers toorganisms usually consideredtobe nonpatho-genic except in patients with predisposing factors, such asimmunosuppressionorindwelling foreign bod-ies. Isolates considered to represent contaminants included: (i) nonpathogens which could not be corre-lated with eithera sourcefor the organism or clinical signs of infectionorsepsis,(ii) nonpathogens withouta sourceinpatients with infection duetopathogens, and (iii) pathogens without asource in patients with no clinicalsigns of infectionorsepsis consistent with that organism.
Isolates classifiedastransientbacteremia included: (i) nonpathogens inpatients with a clearly identifiable
source and route into the bloodstream but without eitherclinicalsigns indicative of seriousorpersistent infection or signs of bloodstream infection by that organism and(ii) pathogens in patients withaclearly identifiable source but without either signs of serious
orpersistent infection orsigns of bloodstream infec-tionby thatorganism. Isolates consideredtorepresent septicemia included: (i) nonpathogens in patients with
anidentifiablesourceandrouteinto thebloodstream, with clinical signs of persistent or serious infection consistent with that organism, and with no other identifiable cause for the clinical manifestation of infection and (ii) pathogens in patients with clinical signs and symptoms consistent with infection of the bloodstreamby thatorganism.
One thousand bloodspecimenswere cultured
by the ISOLATOR and BACTEC techniques
over a 5-monthperiod. Ninety-one yielded
orga-nisms that were clinically significant. From
these positive blood
specimens,97 clinically significant organisms were isolated: 57 by both
bythe ISOLATOR system
bythe BACTEC system
only.Table 1 lists thenumber of
cultured from 10
technique only.The BACTEC media did not
grow any S.
pa-tients),and a Clostridium sp. The ISOLATOR
detect-ed in BACTEC media: a Candida albicans
on February 7, 2020 by guest
COMPARISON OF L-C AND RADIOMETRIC TECHNIQUES 1029
TABLE 1. Numberof clinically significant organisms isolated by one or both techniques
No.of isolates detected by:
Organisms BACTEC ISOLATOR BACTEC and
only only ISOLATOR
Staphylococcusaureus 3 3 13
Staphylococcus epidermidis 0 13 10
Group D streptococci 1 1 5
Beta-hemolytic streptococci, not group A, B, D 1 1 0
Streptococcus mutans 0 0 3
Alpha-hemolytic streptococci 0 1 0
Haemophilusparainfluenzae 1 0 0
Escherichia coli 1 1 5
Enterobacter agglomerans 0 1 0
Enterobacter aerogenes 0 1 2
Enterobacter cloacae 0 0 2
Serratiamarcescens 0 0 2
Klebsiella oxytoca 0 0 1
Pseudomonas aeruginosa 1 0 4
Acinetobacter calcoaceticus 0 0 2
Aeromonashydrophila 0 0 2
Bacteroidesfragilis 0 0 4
Bacteroides spp. 1 0 0
Clostridiumperfringens 3 0 0
Clostridium spp. 1 0 0
Lactobacilluscasei 0 2 0
Cryptococcusneoformans 0 1 0
Candida albicans 0 1 0
Candida krusei 0 0 1
Candidaparapsilosis 0 0 1
Candidaglabrata 0 1 0
initially onlyon the
30°C, andin each case,
significant organisms detected
by the ISOLATORsystem
exception of five
albicans,one C. neofor-mans,
patients receivingantibiotics active
ISOLATORsys-tems, 17 werepresent at <1
ISOLATOR method. Table2
of strains of
organisms isolated by bothsystemsand the numberof CFU per milliliter as determined by the ISOLATOR system.
Duringthe course of this study, nine blood cultures from a total of four
organisms.All nineof these polymi-crobial bacteremic
episodeswere detected by the BACTEC system. Three of these episodes
missedby the ISOLA-TOR system.
Organismsundetected by the ISO-LATOR system were a Clostridium sp., a C.
perfringens strain,an S. aureus strain, and a
Pseudomonassp. The BACTEC systemdidnot
between thetwo sys-tems.The
did the BACTECsystem,
detected16 sooner than
ISOLATORsystem, and 28 were
Forty-six organisms yielded
rapidly by the ISOLATOR
system, and in 36
instances, isolated colonies
.24h sooner by the ISOLATOR system.
by the ISOLATOR
tech-nique. Of these,98
single colonyon oneoffourplates inoculated is equivalent to a concentration of 0.125
CFU/ml.The contamina-tion ratefor the radiometric system was
studyconfirms the study of Dorn et al. (1) that the ISOLATOR technique is a more sensitivemethod of
technique alone,24of 27 had
only17 of 57
on February 7, 2020 by guest
TABLE 2. Numberof clinically significant organisms isolated in cultures positive by both the
ISOLATOR and theBACTEC systems
<1 1-10 11-100 >100
Staphylococcusaure- 4 2 5 2
Staphylococcus epi- 4 3 3
GroupDstreptococci 2 2 1
Escherichia coli 1 4
Enterobacteraero- 1 1
Enterobacter cloacae 2
Serratia marcescens 2
Klebsiella oxytoca 1
Pseudomonas aeru- 4 ginosa
Acinetobacter calco- 1 1
Aeromonas hydro- 2
Candida krusei 1
a Numbers refer to numbers of strains of each
organism isolated per the indicated number of CFU per milliliter.
b CFU per milliliter as determined by the
CFU/ml.Dorn et al. (1) and Kiehn et al. (T. E.
Kiehn,C. Capitolo, A.De La
Cruz, andD. Armstrong,
Abstr.Annu. Meet. Am.
from bacteremia.Although it
organisms isolatedat >1
necessarilyrepresent con-taminants and cannot be
distinguish contaminants from pathogens on the
quantitationlimit the usefulness of the ISOLATOR
technique. Contaminationwith the ISOLATOR
technique persistedthroughout the
despitemeasures to control it: the intro-duction of
cotton-plugged ventingneedles, the inoculation of culture
platesin a laminar-flow hood, the inclusion of blood specimens collected
phlebotomists,and theuseof the
same two trained
the Isolator tubes.
technique,ascompared with conventional bottle systems, isamoresensitive
means to detect yeasts. Bille et al. (J.
Stockman,and G. D.
Antimicrob.Agents Chemother. 22nd,
Miami Beach, Fla., abstr.no.
reported a more
rapiddetection time and an
yieldof yeasts and molds with
(93.6% by L-C versus 55.3% by biphasic brain heart
iso-lates, including a C.
transplantpa-tientseen as an
outpatientforaroutine postsur-gical follow-up.
detected in this
study.Four of these were
Five anaerobeswere not
technique butwere cultured in
the BACTEC media. This difference inthe
fact that the 0.4 ml of concentrate
medium is inoculated with4ml
Clostridiumspp. Dorn et al. (1) showed that
isolated byboth the
bottlemethod they were
study of 7,000 blood cultures, Kiehnet
their two-bottle brothsystem, 13 were
by broth only, and11 were
only (Kiehnet al.,
Annu. Meet. Am.
by broth only,and two
by bothmethods (T. E.
communication).S. aureus and Pseu-domonas strains have been
reportedto be de-tected more
comparisonwith the BACTEC system.
isolates,16 were detected
FiveP. aeruginosa strainswere
byboth the ISOLATOR and BACTEC systems.
only bythe ISOLATOR
(3)did demonstratean increasedisolation of S.
lysis-filtrationin comparison with a
conventional blood culture system. Of
processed byconvention-al and
epidermidiswas isolated from
2.4%in commercial brain heartinfusion bottles and from
on February 7, 2020 by guest
contaminant,and only a
chart review demonstratedthe
signifi-cance of the 13 isolates reported here. Seven of the
represented bacteremiaprobably due to an indwelling intravenous catheter, and
endocarditisor a shuntinfection.
epidermidisisolates was either 0.125 or
exception.These counts are
isolation ofone or two
onlyoneof the four
Isen-berg, who also
compared isolation by the
and BACTEC techniques (2).He
technique. Our results show that 87% (84 of 97)
isolated by the radiometric method.
prohibitsthe use of the system for routine
blood culturesaresubmitted to our labo-ratory
annually.The cost per test
is,of course, based on
workload unitsassigned to blood
assigns4.8 U to each BACTEC bottle and 8.8 U to each ISOLATOR blood culture. At Hartford
Hospital,where 1.15 U is
BACTEC blood culture,a
blindsubculture is not
performed, and6.21 U
is assignedto each
ISOLATOR bloodculture, the cost per blood
De-spite the increasedcost, in cases
does offeranadvantage over the the BACTEC system.
Thisproject was supported by a grant from E. 1. du Pont de Nemours &Co.
Wethank the members of the Hartford Hospital phleboto-my team fortheir assistance and interest in this study.
1. Dorn, G. L., G.A. Land, and G.E. Wilson. 1979. Im-proved blood culture technique based on centrifugation: clinical evaluation. J. Clin. Microbiol. 9:391-396.
2. Isenberg, H. D. 1983.Clinical laboratory comparison of the lysis-centrifugation blood culture technique with radiomet-ricandbrothapproaches, p. 38-54. In A. Balows and A. Sonnenwirth (ed.), Bacteremia: laboratory and clinical aspects. Charles CThomasPublishers, Springfield, Ill.
3. Zierdt, C.H. 1983.Evidence for transientStaphylococcus epidermidis bacteremia in patients and in healthy humans. J. Clin. Microbiol. 17:628-630.