j. Soc.Cosmet.
Chem.,35, 253-263 (August 1984)
Comparison
of the API 20E, Flow,and Miniteksystemsfor
the identification of enteric and nonfermentative bacteria
isolated from cosmetic raw materials
JOHN K. McLAUGHLIN, BARRY D. ZUCKERMAN,
SAUL TENENBAUM, and BARBARA A. WOLF, RevlonResearch
Center,2121 Route27, Edison,NewJersey08818.
Received
February1O, 1984. Presented
at theSCC-SIMMicrobiological
Seminar,New York, December
9, 1981.
Synopsis
Five commercialidentificationsystems,(API 20E, Flow Enteric-Tekand N/F System,and Minitek for
Enterobacteriaceae
andNonfermenters)werecomparedwith conventionalmethodologyin the identification
of 94 gram-negativebacteriaobtainedfrom cosmeticraw materialsand referencesources.The percent
agreement
betweenrapidandconventional
systems
for microbialidentifications
rangedfrom 76-88% and
75-79% for entericand nonfermentative
microorganisms,
respectively.
Regardingindividualbiochemical
tests,the percentagreementbetweenthe rapidandconventional
systems
rangedfrom 90-96% for enterics
and 80-90% for nonfermenters.
Greatestvariationswere found in the accuracylevelsof individualbiochemicaltests,with API and Minitek showingthe greatestnumberof inaccuratetestsfor enterics(254% and 2-51%, respectively)
and Flow showingthe least(2-15%). Flow alsoexhibitedthe smallest
incidenceof inaccuratereactionsfor nonfermenters,2-23%, while API and Minitek showed2-68% and
4-45%, respectively.Although relativelycloselevelsof identificationagreementbetweenrapid and
conventionalmethodswere observed,Flow showedthe highestpercentages
of biochemicalcorrelationsto
conventionalsystemsfor the microorganisms
studied. With regard to microorganisms
isolatedfrom industrialsources,
it hasbeenour experience
that the Flow systems
aremostsatisfactory
for both biochemical
accuracyand efficientuseof manpower.
INTRODUCTION
Increasingmicrobiologicalsurveillanceon raw materialsusedto manufacturecosmetic
productshas resultedin the need for accurateand rapid microbialidentifications.
Industrialmicrobiologylaboratories
havefrequentlyrelieduponthe useof conventional
identificationprocedures.
Thesemethods,althoughconsidered
to be accurate,require
high levelsof expertisedue to complexand time consuminginoculationand interpretation procedures.The introductionof the multimedia test systems,which claim to
providerapid and accurateidentificationsutilizing computerizedtechnology,appeared
to fulfill
this need.
The API 20E (API) is frequentlyacceptedasthe systemof choicefor the identification
of enteric (ENTB) and nonfermentative(NFB) bacteria, and often is used to replace
253
254
JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS
conventionalmethods.Our experiences
with API haveindicateda relativelylow degree
of accuracyfor NFB isolatedfrom cosmeticsources,often requiringconfirmationof
API identificationswith conventionalmethodsor other identificationsystems.Studies
utilizing clinicaland foodisolates(1-4) indicatethat the Flow (FL) and Minitek (MT)
systems
are able to identifymicroorganisms
with a high degreeof accuracy.
However,
few studieshave been reportedevaluatingtheserapid identificationsystemsusing
ENTB and NFB isolated from cosmeticsources.Therefore, we undertookto evaluate
the FL, MT, and API systemswith microorganisms
originallyisolatedfrom-rawmaterials used in cosmeticproducts.Our aim was to selectan identificationsystemor
systemswhich couldcorrelatewith conventional
mediaand provideaccurateand rapid
identifications
MATERIALS
of both ENTB
AND
and NFB.
METHODS
CULTURES
The 94 microorganisms
tested(41 ENTB and 53 NFB) included79 commonlyencountered raw material
isolates and 15 stock reference cultures obtained from the
AmericanType Culture Collection,Flow Laboratories,
New York City Departmentof
Health, and RocheDiagnostics.
The microorganisms,
maintainedon Trypticasesoyagar(BBL, Cockeysville,MD) slants
at 4øC, werecheckedfor purity on blood(BBL) agarplates.One colonyfrom each
platewastransferred
to 10 ml of Trypticasesoybroth(BBL), andincubatedat 25øC
for 18 and48 hr for ENTB andNFB respectively.
One ml fromeachbrothwaspipetted
ontoa Trypticasesoyagarplate and spreadoverthe surfaceof the agarto producea
bacterial
lawn.Theplateswereincubated
at 35øCfor 18 h, andtheresultingmicrobial
growth wasusedto inoculateall conventionalmediaand test systems.
TEST
SYSTEMS
(1) API 20E (AnalytabProducts,
Inc., Plainview,N.Y.). Previousexperiences
in our
handsusing API haveoften resultedin inconsistentand variabletest reactions.These
problemswere alleviatedto somedegreeby increasingthe recommended
inoculumsize
to approximatelya No. 5 MacFarlandstandard.The inoculumwasthen addedto each
strip accordingto the manufacturer'sinstructions.ENTB identificationswere completedat 24 h; NFB identifications
werecheckedat 24 h, reincubated,andcompleted
at 48 h. Microorganisms
wereidentifiedby acceptingthe first choiceof the API Profile
Recognition System.
(2) FlowEnteric-Tek
and N/F System
(FlowLaboratories,
McLean,Va.). The Enteric-Tek
systemprovidesfor the determinationof 14 differentbiochemicalparametersby utilizing a wheel of solid media. The systemswere inoculatedand incubatedaccordingto
the manufacturer'sinstructions.Microorganisms
were identifiedby acceptingthe first
choiceof the Flow Enteric-TekSystemcomputercodebook.
The N/F Systemconsists
of the two tube N/F-Screen,and the Uni-N/F-Tek plate. The
N/F-Screen,composed
of the GNF and 42P tubes,containsfive testsutilizedfor the
identificationof fluorescentpseudomonads.
The Uni-N/F-Tek plate is a multichamberedwheel designedfor the determinationof 12 different biochemicalparameters.
COMPARISON
OF BACTERIAL
IDENTIFICATION
SYSTEMS
255
The N/F-Screensand Uni-N/F-Tek plateswere inoculatedsimultaneously,
and incubatedaccordingto the manufacturer's
instructions.Microorganisms
wereidentifiedby
acceptingthe first choiceof the Flow N/F Systemcomputercodebook.
(3) Minitek (BBL, Cockeysville,
Md.). Both MT systemsusedisksthat havebeenimpregnatedwith the substrates
to betested.Numerousothersubstrate
disksareavailable;
however,the diskschosenfor this studywere thoserequiredby the manufacturerfor
the Enterobacteriaceae
II and Nonfermenternumericalidentificationsystem.Inoculations, incubations,and interpretationswere performedaccordingto the manufacturer's
instructions.
CONVENTIONAL
MEDIA
Eachculturewasidentifiedby conventional
biochemical
tests(5,6). The testsperformed
on ENTB included reactionson triple sugariron agar (H2S production);oxidasefrom
blood(BBL) agar;lysineandornithinedecarboxylase
(LDC, ODC); argininedihydrolase
(ADH); citrate;DNase; esculin;gelatin;malonate;methyl-red;Voges-Proskauer
(VP);
nitrate reduction;O-nitrophenyl-B-D-galactopyranoside
(ONPG); phenylalanine;reactions in SIM medium (H2S and indole production;motility); Christensenurea agar;
and productionof acid in phenolred adonitol, arabinose,cellobiose,dulcitol, glucose,
glycerol, inositol, lactose,maltose,mannitol, melibiose,raffanose,rhamnose,salicin,
sorbitol,trehalose,andxylose.The testsperformedon NFB includedreactions
on triple
sugariron agar (H•S production);oxidasefrom blood(BBL) agar;acetamide;LDC and
ODC; ADH; Cetrimide (Difco, Detroit, MI); citrate; DNase; escuiin;gelatin; growth
in Trypticase
soybroth(BBL)at 42øC;lecithinase;
MacConkey
agar;nitrateandnitrate
reduction;ONPG; penicillin and polymyxinB susceptibility;phenylalanine;PseudomonasF (Difco) agar; Pseudomonas
P (Difco) agar;Pseudosel
(BBL) agar;reactionsin
SIM medium;6.5% NaCI; SSagar;starch;10% lactose(purpleagarbase);Tween80
hydrolysis;Christensenureaagar;VP; and oxidationof OF basalmediumof fructose,
galactose,glucose,lactose,maltose,mannitol, sucrose,and xylose.All media inoculatedfrom the bacteriallawnwereincubatedat 35øCexceptwhereindicated.Oxidase
testswere performedusing Cepti-Seal(Marion Scientific,KansasCity, Mo.) oxidase
reagentat 24 h. The reagentsfor indole, nitrate reduction,and phenylalanine
tests
wereaddedafter 24 h of incubation.After 48 h of incubation,reagentswereaddedfor
DNase, methyl red, VP, and starchhydrolysis.Reactionsnegativeafter 24 h were
observedfor a maximumof 7 dayswith the exceptionof gelatin, which wasobserved
for up to 30 days.
RESULTS
IDENTIFICATION
AGREEMENT
AMONG
METHODS
The agreementsamongthe conventionaland rapid systemsfor the identificationof
ENTB and NFB are shownin TablesI and II. Among ENTB (Table I), FL correctly
identified36 of the 41 microorganisms
tested(88% accuracylevel), the highestof the
threesystems.API and MT exhibited80% and 76% accuracy
levels,respectively,
for
ENTB. Closerlevelsof identificationaccuracywereobservedfor NFB (Table II). API
and FL correctlyidentified42 of the 53 microorganisms
tested(79% accuracy).
MT
wasslightlylower, exhibitinga 75% level of accuracy.
256
JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS
Table
I
AgreementAmongConventional,API, Flow and Minitek Systems
With EntericCultures
No. of StrainsCorrectly
Identifiedto Species
Levela
No. of Strains
Organism
Citrobacter
freundii
Enterobacter
cloacae
(1)•'
Enterobacter
gergoviae
(1)
Tested
API
Flow
Minitek
4
4
8
4
4
3
4
2
6
4
2
4
Escherichia
coli (2)
2
2
1
2
Klebsiella
oxytoca
KlebsMla
pneumoniae
(1)
Morganella
morganii
4
6
1
4
5
1
4
6
1
2
6
1
Proteus
vulgaris(1)
1
1
1
1
Providencia stuartii
1
1
1
1
Providencia
rettgeri(1)
2
2
2
1
Salmonellacholerae-suis
(1)
Salmonellaenteritidis(1)
1
1
1
1
1
1
1
1
Serratialiquefaciens
2
2
2
2
Serratia marcescens
3
1
3
3
1
41
1
33 (80%)
1
36 (88%)
0
31 (76%)
Shigellasonnei
(1)
Total
Without the useof supplementaltests.
Numberin parenthesis
indicates
thenumberof reference
strains
used.
Table
II
AgreementAmongConventional,
API, Flow, andMinitek Systems
With Nonfermentative
Cultures
No. of StrainsCorrectly
No. of
Identified
to Species
Level
a
Strains
Organism
Tested
API
A cinetobacteranitratus
1
Acinetobacter
haemolyticus
Acinetobacter
lwofjS'
Alcaligenes
odorans
1
1
1
Moraxella urethraIls
Pseudomonas
aeruginosa
(2)•'
Pseudomonas
cepacia
(1)
Pseudomonas
fluorescens
(1)
Pseudomonas
maltophilia
(1)
Pseudomonas
pseudoalcaligenes
Pseudomonas
putida
Pseudomonas
putrefaciens
Pseudomonas
stutzeri
Total
Flow
Minitek
0
1
0
0
1
1
0
1
1
0
1
1
1
0
0
0
7
7
7
5
15
15
15
15
6
4
3
7
2
6
3
6
1
5
3
0
7
1
0
5
1
0
5
1
4
53
1
42 (79%)
4
4
42 (79%)
40 (75%)
Without the useof supplementaltests.
Numberin parenthesis
indicates
thenumber
of reference
strains
used.
Comparisons
of the correlationlevelsof the threesystems
biochemical
test resultsto
those obtained with conventional methods are shown in Tables III and IV. FL exhibited
the closestoverallagreementwith conventional
testing,96% for ENTB and 90% for
NFB. The samepattern was reflectedamongthe biochemicaltestswhich all three
COMPARISON
OF BACTERIAL
IDENTIFICATION
Table
SYSTEMS
257
III
Percentagreementof APT, Flow, and Minitek With ConventionalBiochemicalTestsfor
Enteric
Test
APT
ADH
Gelatin
90
93
H2S
Tndole
LDC
Malonate
98
98
90
Cultures
Flow
Minitek
80
100
98
95
95
95
98
90
90
NO3-NO 2
100
ODC
ONPG
98
100
93
98
100
95
98
95
100
Phenylalanine•
Adonitol
Arabinose
100
98
98
98
Glucose
100
100
Tnositol
85
Lactose
Mannitol
Mellibiose
88
98
78
85
93
98
78
Raffinose
98
Rhamnose
98
98
98
Sorbitol
95
95
93
Sucrose
Simmons Citrate
95
83
93
Urea
46
95
93
Voges-Proskauer
98
Total
92%
95
49
71
96%
90%
a API and Flow usetryptophanedeaminasein lieu of phenylalanine.
systemshad in common,with FL having the highestlevel of correlationwith conventional testingfor both groupsof microorganisms.
CAUSES
OF MISIDENTIFICATIONS
Falsepositiveand negativetest reactionsand inconsistencies
in the data baseinterpretation of thesereactionscausedmisidentifications
in all systems.TablesV, VI, and
VII
list the causes of identification
differences between conventional
methods and the
rapid systemsfor both ENTB and NFB.
For API, misinterpretationof the rhamnosetest by the data baseresultedin the
identificationof two strainsofSerratiamarcescens
asS. liquefaciens.
API listedS. marcescens
as98% rhamnose
negativeandS. liquefaciens
as30% rhamnose
positive.Rhamnose
was
negativefor both strains;however,the identificationwas still directedtoward S. liquefaciens.
Among NFB, MoraxellaurethraIlswas identified as Pasturella-Actinobacillus
speciesdue to a negativetryptophanedeaminase(TDA) test. Although API listed all
Moraxellaspeciesas 100% TDA negative, somespeciesare phenylalaninepositive (7)
and resultsof thesetestsare usuallyinterchangeable.
API wasalsonot able to identify
any of the three strainsof Pseudomonas
pseudoalcaligenes
evaluated.Two strains were
identified as Pasturella-Actinobacillus
becausethe data basecould not separatethe two
258
JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS
Table
IV
PercentAgreementof API, Flow, and Minitek With ConventionalBiochemicalTestsfor
Nonfermentative
Test
API
Acetamide
ADH
DNase
91
Minitek
92
91
91
85
42øC
83
Gelatin
77
LDC
94
NO3-NO 2
NO2-N 2
94
87
ODC
9!
OF Glucose
74
OF Lactose
100
96
94
94
100
9!
87
85
32
64
OF Xylose
ONPG
98
Phenylalaninea
Pyocyanin
91
Simmons Citrate
Starch
92
Urea
Indole
Flow
91
Esculin
Fluorescein
OF Maltose
OF Mannitol
OF Sucrose
Cultures
79
9!
70
75
87
89
98
96
89
100
55
100
H2S
Voges-Proskauer
98
49
Total
80%
89
92
77
100
55
100
100
90%
88%
a API and Flow use tryptophanedeaminasein lieu of phenylalanine.
groupsof microorganisms,
asmostof the testreactionswerenegative.The third strain
wasplacedin the otherPseudomonas
spp.group,whichincludedP. pseudoalcaligenes
and
six other microorganisms.
As such, the identificationwasconsideredincorrectdue to
the apparentinability of the systemto speciatethis microorganism
or give a smaller
numberof choices.A strainof ?. stutzeriwasidentifiedasAchromobacter
species
due to
a positiveureasetest. This misidentificationwasa resultof the API data baserequirement that all strainsof ?. stutzeribe 100% ureasenegative.However,?. stutzerihas
beenreportedto be asmuchas 18% ureasepositive(6).
With Flow, identificationdifferences
amongENTB and NFB were due primarily to
falsepositiveand negativereactions.In contrastto the API andMT systems,
onlyone
misidentification
wasdue to an informationdeficiencyin the databaseof the system.
Of the threestrainsof P. pseudoalcaligenes
evaluated,onewasidentifiedasAchromobacter
xyloseoxidans
due to a positivexylosetest. The FL databaserequiredP. pseudoalcaligenes
to be 100% xylosenegative;however,xylosecouldbe 14% positivefor this organism (6).
Misidentifications
obtainedwith MT weredue to a combinationof falsepositiveand
negativebiochemicaltest reactionsand data basedeficiencies.
AmongNFB, Acineto-
COMPARISON
OF BACTERIAL
Table
Causes of Identification
IDENTIFICATION
SYSTEMS
259
V
Differences Between Conventional Methods and API a
Cause of
Conventional
Identification
Identification
API Identification
Differences
Enterobacteriaceae
E. gergoviae
(4)b
E. aerogenes
URE(- ), CIT(- ), SORB(-)
E. gergoviae
(1)
K. pneumoniae
(1)
S. marcescens
(2)
S. liquefaciens
Klebsiellaspecies
S. liquefaciens
LDC ( - ), SORB(+ ), GEL(+ )
CIT(- ), URE(-)
Data base
interpretationof neg.
rhamnose c
A. anitratus(1)
Nonj•rmentative
Bacilli
P. paucimobilis
N2( + ), GLU(- ), MEL(-)
A. haemolyticus
(1)
A. anitratus
Omission
of A. haemolyticus
M. urethralis(1)
Pasturella-
Omissionof TDA ( + )
from data base
Actinobacillus
P. maltophilia
(1)
P. pseudoalcaligenes
(2)
Moraxella from data
basec
P. pseudoalcaligenes
(1)
P. cepacia
PasturellaA ctinobaci
llus
Pseudomonas
species
ODC ( + )
All test reactions
negative
Data basecouldnot
identify organismto
P. putrefaciens
(1)
Pseudomonas
species
H2S(-)
P. stutzeri(2)
P. stutzeri(1)
P. aeruginosa
Achromobacter
species
ADH ( + )
Data baserequired
specieslevel
P. stutzeri to be 100%
URE ( - )c
a First choiceof the systems'spectrumof identifications.
bNumberin parenthesis
indicates
thenumber
of strains
misidentified.
c Informationdeficiencies
in the data baseinterpretationof biochemical
test resultscontributedto or
caused misidentification.
bacteranitratuswasidentified asCardiobacterium
hominis,which illustratedan information
deficiency
in the system's
database.C. hominis
waslistedas 100% indolepositiveand
A. anitratusas 100% indolenegative.Althoughthemicroorganism
wasindolenegative,
it was still identified as C. hominis.Two strainsof P. aeruginosa
were misidentified
becauseof positiveurea tests. The MT data basedid not allow for any urea positive
strainsof P. aeruginosa,
even though positive ureaseproductionas high as 74% has
beenreported(6). One strainof P. maltophilia
wasnot in the MT data baseasa result
of falsepositiveanaerobicdextrosereactionsand positivesucrose
and xylosereactions.
MT required P. maltophiliato be 100% sucrosenegativeand 91% xylosenegative.
However, reportsof studieswith conventionalmethodshavefound sucroseand xylose
to be 92% and 56% positive(6) respectively.
DISCUSSION
Each of the methodsutilized in this study had unique characteristics;
therefore,one
would not expectidentificationsobtainedwith onesystemto correlate100% with those
260
JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS
Table
VI
Causes of Identification Differences Between Conventional Methods and Flow a
Cause of
Conventional
Identification
Identification
Differences
Flow Identification
Enterobacteriaceae
E. cloacae
(1)•'
S.fonticola
LDC (+)
E. cloacae
(1)
S. liquefaciens
LAC (-)
E. gergoviae
(1)
E. gergoviae
(1)
K. pneumoniae
Serratia
fonticola
ODC (-)
SORB( + )
E. coli(1)
Serrati•odorifera
ODC (-)
Nonfer•entative
Bacilli
A. haemolyticus
(1)
A. anitratus
Omission
of
A. haemolyticus
from
data base
M. urethraIls
(1)
P. maltophilia
(3)
P. pseudoalcaligenes
(2)
P. pseudoalcaligenes
(1)
P. fluorescens/putida
P. paucimobilis
P. fluorescens/putida
A. xyloseoxidans
GLU(+ ), XYL(+ ), MALT(+ )
DNA(- ), URE(- ), XYL(+ )
GLU (+), MANN (+),
XYL (+)c
P. putida(1)
P. putida(1)
P. aeruginosa
P. picketti
ACET( + ), 42øC( + )
42øC(-), LAC(-),
P. putrefaciens
(1)
P. dimunta
H2S(-)
MANN
( - ), MALT ( - )
a •irst choiceof the systems'
spectrum
of identifications.
bNumberin parenthesis
indicates
thenumber
of strains
misidentified.
c Informationdeficiencies
in the data baseinterpretationof biochemical
test resultscontributedto or
caused misidentification.
of another.However, a parameterhad to be established
which would determinethe
accuracyof the five systems.The parameterdecideduponwasthat of the systems'
ability to correlateidentifications
with thoseobtainedby conventional
methodswhich
wereaccepted
as correct.The identifications
obtainedusingAPI, FL, and MT were
thencompared
to the conventional
identification,
evaluating
the accuracy
of identification, as well as that of individual biochemicaltests.
Inaccuraciesin microbial identificationswere encounteredin all three systems.These
included aberrant biochemicaltest resultsand inconsistencies
in the information pro-
videdby systemdatabases.The combination
of thesetwo problems
exposed
shortcomingsin eachsystem.The majorityof theproblematic
identifications
obtained
with API
werea resultof falsenegativebiochemical
testreactions.
This mayhaveresultedfrom
the system's
apparentinabilityto detectpositivereactions
amongthe weaklysacchrolytic NFB. The causeof this maybe relatedto a lackof nutritivesubstances
in either
the biochemical substrates or in the inoculum vehicle. Difficulties were also encountered
in the interpretation
of the colorchanges
in the decarboxylase
tests.The manufacturer
requires
redor orangereactions
to be considered
positiveat 24 h, andredonlyat 48
h. Frequently
colorsbetweenorangeandredwereobserved
at 48 h, whichresultedin.
uncertaintyin many of the final identifications.
At best, the interpretationof the
decarboxylase
test colorchanges
wasverysubjective.AmongNFB, neitherAPI, FL,
or MT wereable to differentiateA. haemolyticus
from A. anitratus,as the formerhad
not beenincorporated
into anyof the respective
databases.API, however,is the only
COMPARISON
OF BACTERIAL
Table
Causes of Identification
IDENTIFICATION
SYSTEMS
261
VII
Differences Between Conventional
Methods and Minitek a
Cause of
Conventional
Identification
Minitek
Indentification
Identification
Differences
E nterobacteriaceae
E. cloacae
(1)b
S. liquefaciens
ADH ( - )
E. cloacae
(1)
E. gergoviae
(1)
E. gergoviae
(2)
E. gergoviae
(1)
K. oxytoca
(2)
P. rettgeri(1)
S. sonnei
(1)
Serratiaplymuthica
Enterobacter
aerogenes
Enterobacter
vulneris
Salmonella
arizonae
K. pneumoniae
Klebsiella
ozaenae
Salmonella
paratyphi-A
ADH ( - )
URE ( - )
CIT(- ), ODC(- ), VP(H2S(+ ), ODC( - ), VP( IND (-)
IND( - ), LDC( - ), PA( ONPG ( - )
A. anitratus (1)
Cardiobacterium
hominis
IND (-),
A. haemolyticus
(1)
A. anitratus
Omissionof
A. Haemolyticus
from
M. urethraIls(1)
PA (+),
P. aeruginosa
(1)
Pseudomonas
species
P. stutzeri
P. aeruginosa
(1)
Not in database
d
P. fluorescens
(1)
P. maltophilia(1)
P. aeruginosa
Not in data base
P. pseudoalcaligenes
(3)
P. putida(1)
P. putida (1)
P. putrefaciens
(1)
Moraxellaspecies
P. pseudoalcaligenes
P. stutzeri
A lcaligenes-Pseudomonas
species
Nonfermentative
Bacilli
MAC (-)c
data base
MAC (-)
URE (+)c
URE (+), N2 (+)c
ANADEX
(+), ONPG(+)
ANA DEX ( + ), SUC ( + ),
XYL (+)
ADH ( - )
AER DEX (-)
STAR ( + )
AER DEX (-)
First choiceof the systems'spectrumof identifications.
Numberin parenthesis
indicates
thenumber
of strains
misidentified.
Informationdeficiencyin the systems'
databasecontributedto or causedmisidentification.
Profilenumber
generated
by themicroorganism
wasnotin thesystems'
database.
systemwhich containsgelatin, oneof the primary testsusedto differentiatethesetwo
microorganisms.It was not clear why API chosenot to includeA. haemolyticus
as a
possibleidentificationchoicesincegelatinliquefactionis part of their data base.API
in generalexhibiteda largenumberof biochemical
test reactionsthat yieldedunsatisfactoryresultsand that wereat timesdifficultto interpretand ofteninaccurate.
On
the other hand, few difficulties which affected the outcome of identifications were
encounteredin the use of either FL system.One problemnot relatedto biochemical
or systemaccuracywas the interpretationof the centerwell reactionsin the EntericTek whenboth H2S and TDA reactions
werepositive.FL technicalservicesolvedthis
problemby suggestingthe use of two separateinoculationsat oppositesidesof the
centralwell. This allowedfor separateH2S and TDA readings,and facilitatedfurther
identifications.
Like API, problemswith MT centeredaroundquestionable
biochemical
test reactions and lack of information
in areas of the data base. Positive anaerobic
dextrosereactionsobtainedwith strainsof P. cepacia
causedsomeconcern;however,
MT accounted for this reaction in all instances. The MT technical service informed us
262
JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS
that the positive anaerobicdextrosereactionswere due to oxidation rather than fermentationsincethe wells of the MT plate are not deepenoughto preventoxidation,
evenwith an oil overlay.Consequently,falsepositiveanaerobicdextrosereactionsdid
causethe misidentificationof strainsof P. fluorescens,
P. maltophilia,P. putida,and P.
putrefaciens.
One other test causingproblemsfor both ENTB and NFB was urease.
Positiveureareationsresultedin the identificationerrorof two strainsof P. aeruginosa,
one of which was not in the data base,eventhough the MT urea reactionagreedwith
the conventionaltest for both microorganisms.
Like APT, resultswith MT indicated
its ability to correlatewith conventionalmedia was inferiorto FL.
A final areaevaluatedwas the easeof usefor eachof the systems.This consideration
is of prime importanceto many industriallaboratories
that experiencemanpowerdifficultiesand must rely uponpersonnelotherthan microbiologists
to performmicrobial
identifications.This has developedan acute need for a systemwhich can provide
accurateidentificationsand yet be simpleto use.The two FL systemswerethe simplest
to usein our handsbecausethey are suppliedas ready-to-use
productsthat requireno
further assemblyor specialincubationchambersand requireonly Kovacsas an additionalreagentfor the indoletest. In contrast,APTandMT bothrequirefurtherassembly
prior to useand make useof humidificationchambers.API alsorequiresthe useof six
reagents,while MT requiressix for ENTB and five for NFB biochemicalreactions.FL
offeredother benefitsby not requiringmeasuredamountsof substrates
in the wells,
and the mode of inoculation,a Pasteurpipet, was quick and uncomplicated.API
inoculations
requiredhigherlevelsof experience
with respectto filling cupulesproperly.
Even with experiencedpersonnel,problemswith air bubblesand cupulesthat would
not fill properlywere encountered,which resultedin delaysand userfrustration.MT
inoculationswere madedifficult at times by the useof a pipet gun system.Often the
trigger on the pipet gun would be difficult to pull, and severaltimes the pipet tip
cameapartfrom the pipettor, resultingin spillageandlossof the inoculum.In contrast,
the advancedsimplicity of the 2 FL systemsmade them a clear choicefor the easiest
system to use.
To somedegree,our findingsagreedwith studiesevaluatingAPT, FL, and MT using
clinicalandfoodisolates(2-4,8-10). However,sinceour intentwasto selecta system
or systemsto identify both ENTB and NFB, direct comparisons
to other studieswere
not possible.It wasour opinionthat an industrialmicrobiologylaboratorymust consider both ENTB and NFB when making a choiceof identificationsystems.It was
clearto usthat a choiceof systems
couldnot bemadesolelyon the basisof identification
accuracy.The parameter,althoughinformative,is often misusedand is not alwaysa
true indicationof a system'sability, especiallywhen dealingwith industrialmicroorganisms.This is supportedby the fact that the data basesfor eachof the systems
containinformationderivedprimarilyfrom microorganisms
obtainedfromclinicaland
referencesources.Consequently,the biochemicalprofilesderivedfrom industrialisolatesfrequentlyare not containedin the identificationscheme.Therefore,industrial
laboratoriesmust build their own informationlibrariesto supplementinformation
providedby the manufacturer
of the system.The parameterto whichwe gaveprimary
significance
wasthe correlationbetweenconventionaland systembiochemicaltesting.
Basedon this information,a final choiceof a systemwas made. The relativelylow
correlationof APT, and lack of key tests required for the differentiationof NFB,
contributedto the elimination of APT. It was apparentthat no singlesystemcould
COMPARISON
OF BACTERIAL
IDENTIFICATION
SYSTEMS
263
accuratelyidentify both ENTB and NFB, whichcausedus to favorFL and MT. It was
believedthat due to the low viability of many industrial isolates,and their inability
to grow well on artificial media, the systemof choiceshouldbe one offeringthe best
possibleopportunityfor microbialsurvivalandoptimalexpression
of biochemical
characteristics.This studyillustratedthat the two FL systemsprovidedexcellentlevelsof
biochemicalcorrelationwith conventionalmethodology.We believethat this wasdue
to the utilization of conventionalmedia as the basisfor developingbiochemicalreactions. The use of solid media, as comparedto dehydratedsubstrates,allowedfor the
optimalbiochemicalexpression
of the microorganisms,
thusobtainingthe mostaccurate
identificationpossible.
Therefore,it is the opinionof the authorsthat the FL systems
are the most suitablesinglesystemsfor quality controland researchtesting. It is our
hopethat this informationwill stimulatea re-evaluaionof the microbialidentification
systemsnow in use in industrial laboratories.
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