Use of Enzyme Tests in Characterization and Identification of Aerobic and Facultatively Anaerobic

23 Pages·1998·0.24 MB·English

CLINICALMICROBIOLOGYREVIEWS,Apr.1998,p.318–340 Vol.11,No.2
0893-8512/98/$04.0010
Copyright©1998,AmericanSocietyforMicrobiology
Use of Enzyme Tests in Characterization and Identification of
Aerobic and Facultatively Anaerobic Gram-Positive Cocci
SHOSHANABASCOMB1ANDMAMMADMANAFI2*
31BrambleWalk,LymingtonSO419LW,UnitedKingdom,1andHygieneInstitute,
UniversityofVienna,A-1095Vienna,Austria2
INTRODUCTION.......................................................................................................................................................318
GRAM-POSITIVECOCCI........................................................................................................................................319
Micrococcaceae.........................................................................................................................................................319
Micrococcus...........................................................................................................................................................321
Planococcus...........................................................................................................................................................322
Staphylococcus......................................................................................................................................................322 D
o
Stomatococcus.......................................................................................................................................................325
w
Identificationschemesandkits........................................................................................................................325 n
StreptococcaceaeandRelatedOrganisms.............................................................................................................325 lo
Cellsarrangedinpairsorchains,withnocatalaseactivity........................................................................329 a
d
(i)Enterococcus................................................................................................................................................329 e
(ii)Globicatella.................................................................................................................................................329 d

(iii)Lactococcus...............................................................................................................................................329 fr
o
(iv)Leuconostoc................................................................................................................................................329 m
(v)Streptococcus...............................................................................................................................................330
h
(vi)Vagococcus.................................................................................................................................................330 t
t
Cellsarrangedinpairsortetrads,withnocatalaseactivity.......................................................................330 p
:
(i)Gemella........................................................................................................................................................330 //
c
(ii)Helcococcus................................................................................................................................................331 m
(iii)Pediococcus...............................................................................................................................................331 r
.
(iv)Tetragenococcus.........................................................................................................................................331 a
s
Cellsarrangedinpairsortetrads,withweakcatalaseactivity...................................................................331 m
(i)Aerococcus...................................................................................................................................................331 .
o
(ii)Alloiococcus................................................................................................................................................331 r
g
Identificationschemesandkits........................................................................................................................331 /

GENERALCONSIDERATIONSANDINTERRELATIONSHIPBETWEENTESTS......................................333 o
n
DISCUSSIONANDCONCLUSIONS.....................................................................................................................335
A
ADDENDUM...............................................................................................................................................................335
p
REFERENCES............................................................................................................................................................335 r
il
3
,

INTRODUCTION from2htoseveraldays.Completiontimesfortestsperformed 2
0
directlyonclinicalspecimensarealsovariable. 1
Microorganisms have been classified and identified on the Most growth-dependent tests require at least an overnight 9

basis of a variety of characteristics including morphological, incubation;others,basedontheabilitytoutilizeasinglecar- b
y
growth, tolerance, metabolic, biochemical, and genetic. Re- bon or nitrogen source, may require as long as 7 days. The g
centlytherehasbeenatendencytodeterminedefinitiveclas- moleculargenetictechniquesarestilltime-consumingandless u
sificationandtaxonomicassignmentbynucleicacidhybridiza- amenabletoroutineapplication.Moreover,mostofthetech- es
tion,16SrRNAsequenceanalysis,andothermoleculargenetic niquesavailablenowareforthespecificdetectionofalimited t
techniques. After classification has been established, charac- numberoftaxa.Alternatively,determinationsoftheenzymatic
teristicsareselectedfortheidentificationofunknownisolates. activities of isolates with a variety of synthetic substrates (16,
Commercialkitsbasedonsuchaprocessareavailableforthe 52,124)canbeusedforidentificationandgivesimilarresults
identificationofclinicallyimportantbacteria.Itisessentialto tothoseobtainedbyothercharacterizationmethods.Theen-
realize that for routine identification of isolates from human, zymatic characterization of microorganisms by means of syn-
food, or veterinary specimens, ease of testing and total com- theticsubstratesmakesuseofthefactthatmanyenzymesare
pletiontimearecritical,sincetheaddedvalueofidentification constitutivelypresentoreasilyinducedandrapidlydetectable,
information to the clinical or processing outcome decreases often after incubation times of seconds to 3 h. Thus, identifi-
thelateritbecomesavailable.Completiontimesfortheiden- cationofbacteriabasedonenzymepatternsofferssimpleand
tification of bacteria taken from isolated colonies can vary rapidresults.
Theabilitytodetectspecificenzymesrapidlywithsynthetic
chromogenicorfluorogenicsubstrateshasbeenstudiedexten-
sively(17,35,53,54,110,117,118,150).Testsinvolvingsome
*Correspondingauthor.Mailingaddress:DepartmentofFood-and
Waterhygiene,HygieneInstitute,UniversityofVienna,Kinderspital- of these substrates have been included in commercial kits for
gasse15,A-1095Vienna,Austria.Phone:[phone].Fax:43-1- identificationortaxonomicstudiesofbacterialisolates.
40490-295.E-mail:Mohammad.manafi@univie.ac.at. Thefirstcommercialkitwithtestsforspecificenzymescon-
318
VOL.11,1998 ENZYME TESTS FOR GRAM-POSITIVE COCCI 319
tainedPatho-Tecpaperstrips(172),amethodthatevolvedto speaking,teststhatrequiremorethan2hofincubationand/or
the Micro-ID system (15) for the identification of clinically includeagrowth-supportingmediuminthetestcompartment
importantgram-negativerods,mainlymembersoftheEntero- allow the detection of inducible enzymes. Tests requiring in-
bacteriaceae. The Micro-ID kit included tests for b-galactosi- oculum densities of $2 McFarland standard units frequently
dase,cytochromeoxidase,lysineandornithinedecarboxylases, necessitate an additional overnight incubation to achieve a
tryptophanase, and urease. Other kits in the form of cards, sufficientinoculum.
microtiter trays, or multichamber strips are now available for Studies of the clarification of the taxonomic position of in-
theidentificationofcertaintaxonomicclassesofbacteria.The dividualtaxahaveusedlaboratory-preparedtests(117,119)as
largest number is available for the identification of clinically well as commercial characterization and identification kits.
important aerobic and facultatively anaerobic gram-negative Only the latter contain information on expected results for
bacteria(e.g.,API20E,MicroScanconventionalovernightand specifiedtaxaineachofthekittests.
MicroScan Rapid GN Identification Systems, and Vitek GNI The API ZYM system (API System; bioM´erieux, Paris,
Card).Fewerkitsareavailablefortheidentificationofgram- France) is a semiquantitative micromethod designed for the
positivecocci,staphylococci,streptococci,anaerobiccocci,and detectionof19enzymaticactivities(106,232).TheuseofAPI
yeasts.API20E,MicroScanconventionalovernight,andVitek ZYMfortaxonomicstudiesofavarietyoftaxaofbacteriaand
GNI Card include growth-dependent tests and a few enzyme other prokaryotic as well as eukaryotic organisms has been
tests;ingeneral,theydonotusetheabilityofenzymeteststo tabulated(99).TheuseofAPIenzymeresearchkitsdetecting D
provideresultsrapidly.Aspecificenzymetestforb-galactosi- 20glycosidases,10esterases,57arylamidases,alkalineandacid o
w
dasebasedonutilizationofthesyntheticsubstrateo-nitrophe- phosphatases, and phosphoamidase has been reported (151,
n
nol-b-D-galactopyranoside (ONPG) or substrates with other 153,164,226). lo
syntheticmoietiesisincludedinmostkitsforidentificationof In addition, kits based on growth-dependent tests to deter- a
d
the Enterobacteriaceae. Some kits are completely manual, mine the utilization of amino acids, organic acids, and carbo- e
whereas others offer automation with instruments for all or hydrates are available. The API 50CH (12, 123, 164), 50AA, d

some of the following tasks: inoculation, incubation, determi- and50OA(125)testsarestripbased;theresultsareindicated fr
o
nation of test results, and identification. Dade MicroScan bychangesinthecolorofthepHindicators.TheBiologsystem
m
RapidIdentificationSystemsforgram-negativerodsandgram- (Biolog Inc., Hayward, Calif.) is microtiter tray based and is

h
positivecocciarethefastestsystems,providingidentificationin dependent on the detection of substrate-specific dehydroge-
t
t
2 to 2.5 h by measuring enzymatic activities fluorometrically nases with tetrazolium salt as an indicator (163). The MAST p
:
with a high correct identification rate. Thus, overall rates of IDsystem(MastLaboratoriesLtd.,Bootle,England)provides /
/
c
98.4and92.5%correctidentificationtothespecieslevel,with ameansofdeterminingthemetabolicactivitiesofanumberof m
and without additional tests, respectively, were obtained dur- isolatesbyagarplateandmultipointinoculatortechniques(84, r
.
ing the database development phase of the MicroScan gram- 86, 123, 193). Combinations of these systems permit the de- a
s
negativerapididentificationsystemtype3(RNID3)for4,151 tectionofover340biochemicalreactions.Anumberofstudies m
isolates comprising 138 fermentative and nonfermentative report the use of commercially available characterization kits .
o
gram-negativebacteria(1).ClinicalstudiesoftheRNID3sys- alone or in combination with test batteries prepared in their r
g
temshowed96.8and89.5%correctidentificationfor405fresh ownlaboratories(25,31,63).
/

and247challengeisolates,respectively(18). This review will survey the current classification of aerobic o
n
The commercial identification systems also provide data- andfacultativeanaerobicgram-positivecocciwithemphasison

A
basesofexpectedresults,andanunknownisolateisassignedto the role of rapid enzyme tests in characterization and identi-
p
oneofthetaxainthedatabaseeitherbyusingacodebookor fication. r
byusinganautomatedsystemandcomputer-basedidentifica- il
3
tion. Most of these kits have been designed for clinically im- GRAM-POSITIVECOCCI ,
portant groups of bacteria, as reflected in the taxa, tests, and 2
0
expected results included in the databases. Some may be ap- Theaerobicandfacultativelyanaerobicgram-positivecocci 1
plicable to isolates taken from different environments (143, were originally divided into two families, Micrococcaceae and 9

240). These kits can also be used for characterization of mi- Streptococcaceae,onthebasisofcatalaseactivityandcellmor- b
y
croorganismgroupsotherthanthosespecifiedinthedatabase phology and aggregation. However, this classification may g
(27). Methods for easy extraction and analysis of enzymatic change, since genetic studies have indicated thatStaphylococ- u
activitiesarerequiredforsuchapurpose. cus is more closely related to the Bacillus-Lactobacillus-Strep- es
Commercialidentificationkitshavebeenoptimizedforthe tococcusclusterthantoMicrococcusorStomatococcus(129). t
sets of taxa included in their databases. They require regula-
tory approval and are expected to provide a high level of Micrococcaceae
accurate identification in comparison with an acceptable ref-
erence method. Characterization kits, on the other hand, are The family Micrococcaceae, which includes the aerobic and
designedtoprovideaneasyandreproduciblemethodfortest- facultatively anaerobic gram-positive cocci giving a positive
ing a variety of unknown isolates. They do not provide ex- reactioninthecatalasetest,isnotaphylogeneticallycoherent
pectedresultsandarelessrigorouslyregulated.Theyarevery group.Fourgenera,Micrococcus,Planococcus,Staphylococcus,
usefulintaxonomicstudies,andtheresultsfromsuchstudies andStomatococcus,arerecognized(129).Testsusefulforthe
maybeusedfortheconstructionofidentificationkits.Someof separation of these taxa include oxygen requirement for
thesekitshavebeendesignedspecificallyforthecharacteriza- growth,oxidasetest,motility,NaCltolerance,andsusceptibil-
tionofmicroorganismswithavarietyofchromogenicenzyme itytobacitracin,furazolidone,andlysostaphin.Geneticstudies
substrates.Thesekitsarenotaccompaniedbyadatabase,and have suggested that Micrococcus belongs to the actinomycete
some may have methods for computer analysis of the data group, and the isolates originally identified as species of Mi-
includingclassificationandidentificationofunknownisolates. crococcus have since been placed in five different genera (see
Theyvaryinthecomplementoftestsincluded,thesizeofthe below)(218).
inoculum required, and the incubation period. Generally Staphylococcus is the only genus showing susceptibility to
320 BASCOMB AND MANAFI CLIN.MICROBIOL.REV.
TABLE 1. Comparisonofsystemsandschemesfortheidentificationofgram-positivebacteria
Identificationsystema
IdentifiesMicrococcaceaeandStreptococcaceae IdentifiesMicrococcaceae IdentifiesStreptococcaceae
Characteristicand
testtype API Geary Kloosand API Rapid API
MS MSRapid Pasco Staph- RapID
VitekGPI Staph etal. Bannerman Staph ID32 20
GP GP GPID Zym STR
ID (86) (129) ID32 Strep Strep
Characteristics
Inoculumdensity 0.5 0.5 0.5–1 0.5 3 0.5 2 1 4 4
(McFarland)
Inoculumvol(ml) 3 6.5 1.8 3 2 2–3 3 1 2–3 2
No.oftestsincluded 26 34 29 18 10 15 36 26 10 14 32 20
No.ofadditionaltests 2 1 3 2 1 1 0 1 4 2 3 2
No.ofreagents 5 0 0 3 1 0 11 6 3 1 5 5
No.oftaxaincluded 48 50 44 26 17 12 36 32 26 52 48
Storagetemp(°C)b RT 2–8 2–8 2–8 2–8 2–8 2–8 2–8
Mediumforinoculum TSA15%SB TSA15%SB BHI V SBA D
sourceb o
Incubationperiod(h) 16–42 2 4–15 16–20 5 18–24 18–72 24 18–24 4 4 4–24 w
n
Testtype(no.oftests) lo
Glycosidases/phosphatases 4 12 0 3 4 1 4 3 4 4 8 4 a
d
Formationofacid 10 11 18 6 4 10 12 14 2 4 17 10 e
Peptidases 1 10 0 1 0 0 2 2 1 4 3 2 d
Hydrolases 1 0 1 1 0 2 5 1 0 1 1 2 f
r
Othermetabolictests 4 1 2 3 2 1 7 5 3 1 3 2 o
m
Growthandtolerance 6 0 8 4 0 1 6 1 0 0 0
tests h
Mandatoryadditional C,H C C,Cg,H C,H C C C G C,H C,H,P C,H tt
testsc p
:
/
/
aMSGP,MicroScanConventionalGram-PositivePanel;MSRapidGP,MicroScanRapidGram-PositivePanel;VitekGPI,VitekGramPositiveIdentificationCard; c
m
PascoGPID,PascoGram-PositiveID;APIStaphID,APISTAPHIDENT;Gearyetal.(86),basedonmultipointinoculationofagarplates.
bRT,roomtemperature;TSA15%SB,Trypticasesoyagarplus5%sheepblood;BHI,brainheartinfusionbroth;V,variousmedia;SBA,sheepbloodagar. r.
cC,catalase;Cg,coagulase;H,hemolysis;G,growthandtoleranceteststhatneedtobedoneseparatelyfromthekit;P,pigmentation. a
s
m
.
o
r
lysostaphin, but some exceptions occur among species of Mi- appeared recently (129). Some staphylococci are human and g
/
crococcus and Staphylococcus. The methodology and useful- animal opportunistic pathogens that, under certain circum- o
nessofthelysostaphintestfordifferentiationbetweenMicro- stances,aremajorcausesofmortalityandmorbidity.Because n

coccus and Staphylococcus have been discussed (10, 83, 100, of the ability of members of the genus Staphylococcus to ac- A
p
133,134,138,178,184,207,210,253).Lysostaphinisaprotein quire resistance to many antimicrobial agents (e.g., strains of r
preparation derived from culture filtrates of “Staphylococcus methicillin-resistant S. aureus (MRSA), they can cause major il
3
staphylolyticus,” which contains three enzymes capable of af- clinical and epidemiological problems in hospitals (30), and ,

fecting the bacterial cell wall: a glycyl-glycine endopeptidase, their presence is monitored carefully. Schemes for the differ- 2
an endo-b-N-acetylglucosaminidase, and an N-acetylmu- entiation of members of the family have been published (12, 01
ramoyl-L-alanineamidase(32,215).Theendopeptidaseisthe 129, 194). Kits available for the identification of this group 9
componentthatlysesthecellwallofStaphylococcusaureusby includeAPISTAPH-IDENT,Staph-TRAC,APIID32Staph, b
y
hydrolyzing the polyglycine of the pentapeptide bridge be- RAPiDECStaph,andVitekGPICard(bioM´erieux-VitekInc.
g
tweenglycopeptidechainsofthestaphylococcalcellwall.Sloan Hazelwood, Mo., and bioM´erieux S.A., Marcy l’Etoile, u
et al. (215) have shown that the endopeptidase is capable of France);GPMicroplatetestpanel(Biolog);MicroScanPosID e
s
catalyzingbothhydrolysisandtranspeptidationreactionswhen and Combo, and Rapid Pos ID and Combo panels (Dade t
actingonglycylpeptides.Theusefulnessofthetestdependson InternationalInc.,WestSacramento,Calif.);PascoGram-Pos-
the purity of the enzyme preparation, the storage of the en- itiveID(Pasco,WheatRidge,Colo.);andStaph-Zym(Rosco,
zyme,thelysostaphinconcentration(184),andthetestproce- Tastrop,Denmark).Thegeneralrequirementsofthekitsand
dure. When susceptibility is tested by observation of growth thetestsincludedarelistedinTables1to6,whichalsocontain
inhibitionzones,italsodependsonthemediumonwhichthe somepublishedidentificationschemesforthesetaxa.Inaddi-
bacteria are grown (144). Susceptibility also depends on the tiontotheschemeslistedinthetables,Wattsetal.(238)have
cell wall amino acids of the bacterial species; species that testedtheAPI20GP(bioM´erieux-Vitek)kit,whichconsistsof
containserineintheinterpeptidebridge,suchasS.saprophy- 20microcupulescontainingdehydratedsubstratesfortheiden-
ticus, S. haemolyticus, and S. hominis, are less susceptible. In- tificationofstaphylococciandgroupDstreptococci.Itincludes
terestingly, lytic agents such as lysostaphin are produced by the10Staph-Identtestsand10testsselectedfromtheAPI20S
membersofthegenusStaphylococcusonly. streptococcal identification system, but it has provided only
OfthegeneraincludedinthefamilyMicrococcaceae,Staph- 56.10% correct identification. STAPHYtest (Lachema, Brno,
ylococcusisthemostclinicallyimportantandcontains32spe- CzechRepublic),amicrotiter-basedsystemwithninetestsand
cies,mostofwhichliveontheskinandotherexternalsurfaces with eight compartments per isolate for identification of Mi-
of animals (129). A comprehensive review of the genera Mi- crococcus, Stomatococcus, and Staphylococcus, has been de-
crococcusandStaphylococcusandtheirclinicalsignificancehas scribed (211). A system described by Rhoden et al. (194) is
VOL.11,1998 ENZYME TESTS FOR GRAM-POSITIVE COCCI 321
TABLE 2. Comparisonofglycosidaseandphosphatasetestsusedinsystemsforidentificationofgram-positivebacteria
Identificationsystema
IdentifiesMicrococcaceaeand
IdentifiesMicrococcaceae IdentifiesStreptococcaceae
Streptococcaceae
Enzyme
MS API Geary Kloosand API Rapid
MS Vitek Pasco Staph- RapID API20
Rapid Staph etal. Bannerman Staph ID32
GP GPI GPID Zym STR Strep
GP ID (86) (129) ID32 Strep
a-L-Arabinosidase MEU
N-Acetyl-b-D-glucosaminidase MEU PNP 1
N-Acetyl-b-D-galactosaminidase MEU
b-D-Cellobiosidase MEU
b-D-N9,N9-Diacetylchitobiosidase MEU
a-D-Galactosidase MEU PNP 1 6B
b-D-Galactosidase PNP MEU PNP PNP 1 1 1,1b bN
a-D-Glucosidase MEU PNP PNP
bbb---DDD---GGMllauunccnouosriosdindaiasdesaese PNP MMMEEEUUU PNP PPNNPP PPNNPP 1 11 111 AS Dow
Phosphatase PNP,I MEU PNP PNP PNP PNP 1 1 PNP 1 bN n
lo
No.oftests 4 12 0 3 4 1 4 3 4 4 8 4 a
d
aForanexplanationoftheidentificationsystems,seeTable1,footnotea.Syntheticmoietyofsubstrates:AS,naphtholAS-BI;6B,6-bromo-b-naphthol;I,indoxyl; e
bN,b-naphthol;PNP,p-nitrophenol;1,proprietarysyntheticmoiety. d
bTwosubstrateswithdifferentproprietarysyntheticmoieties. fr
o
m

h
t
t
based on 6 screening, 18 primary, and 11 confirmatory tests; have been described (129). Kocur et al. (136) and Baldellon p
the tests used are similar to those described by Kloos and andM´egraud(12)examinedstrainsfrommembersoftheMi- :/
/
Bannerman (129). Eight enzyme tests are included in this crococcaceaebyusingavarietyofAPIteststrips.Kocuretal. cm
scheme,whichreportsresultsandassignsisolatestoataxonby (136)concludedthatMicrococcusspeciesproduceavarietyof r
.
usingasix-digitnumericalcode.TheRAPiDECStaphsystem aminopeptidases. Substrates for separation within the genus a
is based on detection of the activities of three enzymes: “au- Micrococcusincludedhydroxyprolyl,glycyl-prolyl,aspartyl,and sm
rease”,b-galactosidase,andalkalinephosphatase;itrequiresa tyrosyl-seryl conjugates of naphthylamine. M. kristinae, M. lu- .
o
veryheavyinoculumandcanidentifyS.aureus,S.epidermidis, teus, M. varians, M. nishinomiyaensis, M. sedentarius, and M. r
S. saprophyticus. Additional tests are required to differentiate lylae were included in the studies by Baldellon and M´egraud g
/
between S. intermedius, S. xylosus, and the remaining Staphy- (12).Theyshowedthatmostspeciesdidnotexhibitproduction o
lococcusspecies. of acid from the 49 carbohydrates included in API 50CH kit. n

Thenumberoftaxaidentifiedbythesekitsvariesfrom5by TheexceptionswereM.kristinaeandM.varians,whichshowed A
p
RAPiDEC Staph to 50 by MicroScan Rapid Pos ID panels. mostlyfaintactivityaftera24-hincubationwithasmallnum- r
Some of the kits are specific to the Micrococcaceae or just to berofsugars.Acidphosphatase,phosphoamidase,anda-glu- il
3
staphylococci,whereasothersareintendedforidentificationof cosidase activities were present in most Micrococcus species. ,

both staphylococci and streptococci. Tables 1—6 show that Arylamidaseactivitieswerefoundinmostspeciesandcouldbe 2
0
testsfortheformationofacidfromavarietyofsaccharidesare usedtoseparateMicrococcusfromtheothergeneraaswellas 1
usedmostfrequentlyfollowedbytestsforglycosidases,hydro- todifferentiateamongsomeMicrococcusspecies. 9

lases,andpeptidases.Thenumberofenzymetestsincludedin Recently, studies of fatty acid and mycolic acid patterns, b
y
each of these varies from 2 in the Vitek GPI Card to 23 in peptidoglycantype,and16SrDNAanalysisofthetypestrains
g
MicroScanRapidPosIDpanels.Resistancetocertainantimi- of species of Micrococcus, Stomatococcus, Arthrobacter, and u
crobialagentsalsoplaysanimportantroleindifferentiationof related actinomycetes but not species of Staphylococcus have e
s
theseorganisms.TheMicroScanRapidPosIdentificationsys- shown that the genus Micrococcus is heterogeneous. The iso- t
temistheonlyonerelyingonenzymetestsandacidformation latespreviouslyidentifiedasspeciesofMicrococcushavebeen
detectedfluorometricallyandprovidingidentificationofmost placedinfivedifferentgenera:Dermacoccus,Kocuria,Kytococ-
clinically significant staphylococci, enterococci, and strepto- cus, Micrococcus, and Nesterenkonia (218). Thus, M. seden-
cocciin2h. tariusisnowDermacoccussedentarius;M.kristinae,M.roseus,
Micrococcus. The members of the genus Micrococcus differ and M. varians have been removed to the genus Kocuria; M.
fromthoseofStaphylococcusbybeingobligateaerobes,witha nishinomiyaensishasbeenmovedtothegenusKytococcus;and
G1Ccontentof63to73mol%,containingcytochromesa,b, M. halobius has been moved to the genus Nesterenkonia. The
c,andd,lackingteichoicacidsintheircellwalls,lackingglycine genus Micrococcus now contains only two species, M. luteus
intheinterpeptidebridgeoftheircellwalls,beingresistantto andM.lylae.
furazolidone,andbeingsusceptibletobacitracin.Moststrains Of the commercial kits available for the identification of
arealsoresistanttolysostaphin(129);susceptiblestrainsofM. gram-positive cocci, the API STAPH-IDENT database in-
luteushavebeendescribed.Lyticactivitytowardstaphylococci, cludes one entry “Micrococcus sp.,” with an additional test
demonstratedbymembersofthegenusStaphylococcus,isnot tablefordifferentiationamongM.luteus,M.lylae,M.varians,
acharacteristicof98.5%ofmicrococci(205). M. kristiniae, and M. sedentarius; the database of MicroScan
Previously, the genus Micrococcus contained nine species, conventional overnight gram-positive aerobic panels includes
and tests that may help in differentiation between the species one entry “Micrococcus sp.”; the database of MicroScan 2-h
322 BASCOMB AND MANAFI CLIN.MICROBIOL.REV.
TABLE 3. Comparisonoftestsdetectingacidformationinsystemsforidentificationofgram-positivebacteria
Identificationsystema
IdentifiesMicrococcaceaeand
IdentifiesMicrococcaceae IdentifiesStreptococcaceae
Streptococcaceae
Substrate
MS API Geary Kloosand API Rapid
MS Vitek Pasco Staph- RapID API20
Rapid Staph etal. Bannerman Staph ID32
GP GPI GPID Zym STR Strep
GP ID (86) (129) ID32 Strep
n-Acetylglucosamine 1 1 1
Arabinose 1 1 1 1 1 1 1
Arabitol 1
Cellobiose 1 1 1 1 1 1
Cyclodextrin 1
Fructose 1
Glucose 1 1 1
Glycerol 1
Glycogen 1 1
Hemicellulose 1 D
Inulin 1 1 1 1 ow
Lactose 1 1 1 1 1 1 1 1 1 n
Maltose 1 1 1 1 1 1 lo
Mannitol 1 1 1 1 1 1 1 1 1 1 1 a
Mannose 1 1 1 1 1 d
e
Melezitose 1 1 d
Melibiose 1 1 1 f
Methyl-b-D-glucopyranoside 1 ro
Pullulan 1 1 m
Pyruvate 1 1 h
Raffinose 1 1 1 1 1 1 1 1 t
t
Ribose 1 1 1 1 1 1 1 p
Salicin 1 1 1 ://
Sorbitol 1 1 1 1 1 cm
Starch 1
r
Sucrose 1 1 1 1 1 1 1 .a
Tagatase 1 s
Trehalose 1 1 1 1 1 1 1 1 1 1 m
Turanose 1 1 .o
Xylose 1 1 1 rg
/

No.oftests 10 11 18 6 4 10 12 14 2 4 17 10 o
n

aForanexplanationoftheidentificationsystems,seeTable1,footnotea. A
p
r
il
3
,

Rapidgram-positiveaerobicpanelscontainsthreeentries:M. cocci(205).Lackoflyticactivityisconfinedtoasmallnumber 2
0
kristinae, M. roseus, and “Micrococcus sp.,” which covers M. ofisolatesofS.xylosus(205,228). 1
agilis, M. luteus, M. lylae, M. nishinomiyaensis, M. sedentarius, Thetaxonomyofthegenushasbeenstudiedextensively(8, 9

and M. varians. A table of additional tests useful for the dif- 9,86,94,128,129,132).Aself-learningschemecombining35 b
y
ferentiationofthesespeciesisprovided.Testsusefulforsep- biochemical tests with whole-cell fatty acid analysis for the
g
aration in this group include pigmentation, aerobic acid pro- identificationofstaphylococcihasbeendescribed(21).Inthis u
ductionfromglucose,andnitratereductase.Noneofthesekits scheme, isolates were compared to the type strain of each of e
s
reflects the new taxonomy of the genus described by Stack- the 35 taxa for acceptance into the database. The results for t
ebrandtetal.(218). theacceptableisolateswereusedtogenerateafattyacidpro-
Planococcus. Members of the genus Planococcus are cocci file library and a biochemical test table to provide a practical
arranged in pairs or tetrads. They are positive in the catalase systemforidentification.Bothareupdatedwhennewaccept-
andbenzidinetestsandhaveaG1Ccontentof39to52mol%. ableisolatesareaddedtothedatabase.
TheyresembleMicrococcusspp.inbeingstrictaerobes,lacking On the basis of the test for coagulase, the enzyme causing
theabilitytoproducebacteriallyticagentssuchaslysostaphin, coagulation of human and rabbit serum, the genus was origi-
and showing resistance to lysostaphin (205, 228). They can nallydividedintothecoagulase-positivespeciesS.aureusand
tolerate12%NaClandaremotile. coagulase-negative staphylococci (CoNS). Determination of
Staphylococcus. The members of the genus Staphylococcus coagulase activity is still the most frequently used test for the
differfromthoseofMicrococcusbybeingfacultativeanaerobes identificationofS.aureus.
withaG1Ccontentof30to39mol%,containingcytochromes Anumberofmethodsareavailablefordetectingcoagulase.
a and b, containing peptidoglycan and teichoic acids in their Inthecommonlyusedtubecoagulasemethodforfreecoagu-
cellwallswitholigoglycinepeptidesintheinterpeptidebridge lase,overnightbrothculturesareincubatedwithdilutedrabbit
of their cell walls, and being susceptible to furazolidone and plasmafor4hat37°Candthenincubatedovernightatroom
resistant to bacitracin; most strains are susceptible to lyso- temperature; any clotting is deemed positive. In the slide co-
staphin(129).Lyticactivityisproducedby99.5%ofstaphylo- agulase test for bound coagulase, a very heavy suspension of
VOL.11,1998 ENZYME TESTS FOR GRAM-POSITIVE COCCI 323
TABLE 4. Comparisonofpeptidase/proteasetestsusedinsystemsforidentificationofgram-positivebacteria
Identificationsystema
IdentifiesMicrococcaceaeand
IdentifiesMicrococcaceae IdentifiesStreptococcaceae
Streptococcaceae
Substratespecificity
MS API Geary Kloosand API Rapid
MS Vitek Pasco Staph- RapID API20
Rapid Staph etal. Bannerman Staph ID32
GP GPI GPID Zym STR Strep
GP ID (86) (129) ID32 Strep
L-Alanine AMC
Alanyl-phenylalanine-proline 1
L-Arginine AMC bNA bNA
L-Citrulline AMC
a-L-Glutamicacid AMC
Glycyl-tryptophan 1
Hydroxyproline bNA
Leucine AMC bNA
L-Lysine AMC bNA D
Methionine AMC
o
L-Phenylalanine AMC w
L-Pyroglutamicacid AMC bNA 1 n
L-Pyrrolidone bNA bNA bNA 1 bNA bNA lo
L-Tyrosine AMC bNA a
d
e
No.oftests 1 10 0 1 0 0 2 2 1 4 3 2 d

aForanexplanationoftheidentificationsystems,seeTable1,footnotea.AMC,7-amido-4-methylcoumarin;bNA,b-naphthylamine;1,proprietarysyntheticmoiety. fro
m

h
cells is placed on a microscope slide and a loopful of rabbit false-positive results in the slide coagulase test were reported tt
p
plasma is added; a positive reaction is indicated by clumping for9of10and7of10strainsofS.lugdunensisandS.schleiferi, :
/
within10s.Theresultsofthetubeandslidecoagulationtests respectively(85).Testsbasedonrapidlatexagglutinationand /c
arenotalwaysidentical.False-negativeresultsinthetubeand hemagglutination to detect clumping factor or protein A are m
slide coagulase tests have been reported for 8 and 17 strains, also used extensively (79, 131); however, Fournier et al. (78) r.
a
respectively, of 87 strains of S. aureus. In the same study, and Mathieu and Picard (156) showed that they are not reli- s
m
.
o
TABLE 5. Comparisonofmiscellaneoustestsusedinsystemsforidentificationofgram-positivebacteria rg
/

Identificationsystema o
n
IdentifiesMicrococcaceaeand IdentifiesMicrococcaceae IdentifiesStreptococcaceae A
Characteristic Streptococcaceae p
r
MS RMapSid Vitek Pasco SAtaPpIh Geteaarl.y BKalnonoesramnadn SAtaPpIh Staph- RapID RIDap3i2d API20 il 3
GP GP GPI GPID ID (86) (129) ID32 Zym STR Strep Strep ,
2
0
Otherhydrolases 1
40%bileesculin 1 9
DNase 1 b
Heat-stablenuclease 1 y
Esculin 1 1 1 1 1 1 g
Hemolysins 1 1 1 u
e
Hippurate 1 1 s
Staphylocoagulase 1 t
Clumpingfactor 1
No.oftests 1 0 1 2 0 2 5 1 0 1 1 1
Othermetabolictests
Argininedihydrolase 1 1 1 1 1 1 1 1 1 1
Ornithinedecarboxylase 1 1
Catalase 1 1
Nitratereductase 1 1 1 1
Oxidase 1
Productionofacetoin 1 1 1 1 1 1
(VP)
Urease 1 1 1 1 1 1 1 1 1 1
No.oftests 4 1 2 4 2 1 7 5 3 1 3 2
aForanexplanationoftheidentificationsystems,seeTable1,footnotea.
324 BASCOMB AND MANAFI CLIN.MICROBIOL.REV.
TABLE 6. Comparisonofmiscellaneousgrowthandtolerancetestsusedinsystemsforidentificationofgram-positivebacteria
Identificationsystema
IdentifiesMicrococcaceaeand
IdentifiesMicrococcaceae IdentifiesStreptococcaceae
Streptococcaceae
Characteristic
MS API Geary Kloosand API Rapid
MS Vitek Pasco Staph- RapID API20
Rapid Staph etal. Bannerman Staph ID32
GP GPI GPID Zym STR Strep
GP ID (86) (129) ID32 Strep
Colonysize 1
Colonypigmentation 1
Aerobicgrowth 1
Anaerobicgrowth 1
Growthinpresenceof:
Bacitracin 1 1 1
10%bile 1
40%bile 1
Colistin(polymyxin) 1
Crystalviolet 1 D
Deferoxamine 1 ow
Furazolidone 1 n
Micrococcusscreen 1 lo
(Bacitracin) a
Novobiocin 1 1 1 1 1 1 1 d
e
Optochin 1 1 1 d
PolymyxinB 1 f
r
Sodiumchloride(%) 6.5 6 6.5 o
Tetrazoliumred 1 m
Peptonebase 1 h
t
t
No.oftests 6 0 8 4 0 1 6 1 4 0 0 0 p
:
/
/
aForanexplanationoftheidentificationsystems,seeTable1,footnotea. c
m
r
.
a
s
able enough for identification of MRSA isolates, because drolyze the fluorogenic substrate Boc-Val-Pro-Arg-4-methyl- m
strainspossessingcapsularserotype5antigendidnotreactand coumaryl-7-amide.Bulandaetal.(36)usedD-Phe-Pro-Arg-b- .
o
thisserotypeispredominantamongoxacillin-resistantisolates. naphthylamidefordirectdetectionofS.aureus.Thesubstrate r
g
A new latex reagent, Pastorex Staph-Plus (Sanofi Diagnostic profile suggests arginine-specific endopeptidase activity. The
/

Pasteur), consisting of a mixture of latex particles for the de- identity of the amino acid adjacent to the arginine is also o
tection of fibrinogen-binding protein (clumping factor), pro- importantsinceBz-Phe-Val-Arg-p-nitroanilidewasnothydro- n

teinA,andS.aureusstereotypes5and8,hasdemonstrateda lyzedbythecomplex,suggestingthataprolinebutnotavaline A
p
95.1% sensitivity for MRSA isolates (79). Comparison of five isrequiredinthatposition.ThereactionwithChromozym-TH r
agglutination tests (247) has shown sensitivities of 98.9 to wasdoneatpH8.4,andthatwiththefluorogenicsubstratewas il
3
99.6%andspecificitiesof93.9to99.9%. doneatpH7.5.Bulandaetal.(36)confirmedthattheenzyme ,

Positiveresultsinthetubeorslidecoagulasetestshavebeen isliberatedintothemediumintheearlylogarithmicphase(3 2
0
foundforotherStaphylococcusspecies,mainlyS.intermedius, to6h),andthatitsproductiondependsonaerationandarich 1
S.hyicus,andS.lugdenensis(85,189).Vandeneschetal.(225) growthmediumsuchasbrainheartinfusion. 9

demonstratedthatbothsubspeciesofS.schleifericanpromote RausandLove(189)comparedtheactivitiesofstaphyloco- b
y
clotting of rabbit plasma in the standard tube test for coagu- agulases from S. aureus and S. intermedius by using Chro-
g
lase. Positive coagulase activity was reported for S. aureus, S. mozym-TH and concluded that S. intermedius staphylocoagu- u
delphini,S.hyicus,S.intermedius,andS.schleiferi(21). lase resembled S. aureus staphylocoagulase in its rate and e
s
Hemkeretal.(101)providedabetterunderstandingofthe modeofactiononprothrombinbutthattheenzymewaspro- t
staphylococcal clotting mechanism. They have shown that ducedinlesseramountsintheformerspecies.Theyconcluded
staphylocoagulase activates the proenzyme prothrombin in a thatChromozym-THcanbeusedtodetecttheenzymeinboth
stoichiometricreactionbetweenonemoleculeofprothrombin species but that accurate detection of the activity of S. inter-
andonemoleculeofstaphylocoagulase.Engelsetal.(66)sug- medius requires a longer incubation period or preconcentra-
gested that staphylocoagulase could be detected rapidly and tion of the extracellular proteins. In further studies,S. aureus
directly with the chromogenic substrate Chromozym-TH and S. intermedius isolates from humans and other mammals
(Boehringer)fortheroutineidentificationofS.aureus.Morita showeddifferencesintheaffinityofstaphylocoagulasetoward
et al. (168) characterized the enzyme staphylocoagulase of prothrombin of bovine or human origin and in its activity on
S. aureus as an extracellular protein that reacts with pro- human, bovine, and equine fibrinogen. The studies suggested
thrombin in human plasma to form an active molecular com- that the enzymes had structural differences (190). Production
plex which can convert fibrinogen to a fibrin clot and shows of acetoin and acid from maltose and the presence of hyal-
amidase activities. They showed that the staphylocoagulase- uronidaseactivityhavebeenrecommendedasthebesttestsfor
prothrombin complex can hydrolyze the chromogenic sub- differentiation between S. aureus and S. intermedius, with the
strates tosyl-Gly-Pro-Arg-p-nitroanilide (Chromozym-TH), former being positive in all three tests (188). A proprietary
Z-Gly-Pro-Arg-p-nitroanilide, H-D-Phe-Pip-Arg-p-nitroani- fluorogenicsubstratedriedwithprothrombinfordetectionof
lide, and tosyl-arginine-methyl ester (TAME); it can also hy- “aurease”(coagulaseactivityofS.aureus)isusedinthecom-
VOL.11,1998 ENZYME TESTS FOR GRAM-POSITIVE COCCI 325
mercially available RAPiDEC Staph kit. Enzymatic activity is alsolisted.Notincludedinthetableareearlydevelopmentsof
detected visually after a 2-h incubation of a no. 4 McFarland the API systems that have been described (26, 33), studies of
standardinoculum(85,112,165). Staph-TRAC(11)whenusedinconjunctionwithbiochemical
Other enzyme tests, e.g., thermostable nuclease (137, 192), andsusceptibilitytests,andSTAPH-IDENT(179)forcharac-
havealsobeenusedfortheseparationofS.aureusfromother terizationoftaxa.Inaddition,reportsontheidentificationof
groupsofstaphylococci.SincesomeCoNSspeciesareinvolved gram-positive cocci have shown the following percent correct
in human disease and a number of species are also likely to identifications: Pasco Gram-Positive ID (104), 90%; MS GP
develop resistance to antibiotics, interest in the identification panels (104), 87.0%; MS Rapid ID panels (19, 38, 92, 216),
of members of the CoNS group to the species level has in- 95.8, 96.2, 95.7, and 93.0%. MS Rapid ID panels have shown
creased. A review of the CoNS group has been published by 90.0and87.0%correctidentificationwithisolatestakenfrom
KloosandBannerman(128).Thetaxonomyofthisgroupcan sheepandhorsebloodagar,respectively(159).
be difficult, since some morphological characteristics of the Thelevelsofaccuracyofidentificationwiththedifferentkits
coloniesofdifferentspeciesrequireupto4daystoappearand orsystemsvaryfrom38%forMSRapidGPinidentificationof
sincethenumberofseparatingtestsissmall(129).Theability non-S.epidermidisCoNSbloodisolates(241)to100%forthe
to differentiate between virulent and avirulent isolates of the same system in identification of S. aureus without additional
same species on the basis of any characteristic has been diffi- tests (221) (Table 7). Overall percent accuracies depend as
cult.Inthiscontext,thefindingofKedzia(121)thatthephos- muchonthemixtureofisolatestestedasonthesystemused; D
phatase activities of virulent strains of S. aureus were higher studies quite often contain too many isolates of the common o
w
thanthoseofavirulentstrainsisinteresting. species,S.aureusandS.epidermidis,andtoofewisolatesofthe
n
TheenzymesimportantinidentificationoftheCoNSgroup lesscommonspecies.Itisthereforedifficulttoassesstheability lo
are b-galactosidase, b-glucosidase (245), b-glucuronidase, ofasystemtoidentifyataxononthebasisoftestingofoneor a
phosphatase, urease, hydrolysis of esculin (involving b-gluco- twoisolates.Mostsystemsarefairlysuccessfulindifferentiat- de
sidase), and utilization of arginine. Thus, most staphylococci ingS.aureus,S.epidermidis,andS.saprophyticus(13,130).The d

arepositivefortheureasetestbutS.arlettae,S.auricularis,S. identification of the less common species is more variable. fr
o
haemolyticus, S. lentus, S. schleiferi, and S. sciuri are negative. IdentificationofsomeisolatesofS.hominisandS.warnerihas
m
OnlyS.gallinarum,S.lentus,andS.sciuricanhydrolyzeesculin proventobeproblematicwithmostoftheabovesystems(13,

andarepositivefortheb-glucosidasetest.Furtherdetailscan 107, 108, 122, 192, 239). The MS Rapid GP system, which h
t
t
be found in the result tables of the various identification sys- requires2hforincubation,providesalevelofaccurateiden- p
tems. Oberhofer (173) found that all 26 strains of S. haemo- tificationsimilartoMSGP,VitekGPI,andStaphID32,which :/
/
lyticus,thesinglestrainofS.intermedius,and2of7S.warneri require 4 to 24 h of incubation (Table 7). Staph ID 32, Mi- cm
strains tested were positive in the pyrrolidonyl-arylamidase croScan conventional overnight panels, and the Pasco GP ID r
(PYR)test,while65isolatesofS.epidermidis,7isolatesofS. systemgenerallydonottakeadvantageoftheabilityofenzyme .a
hominis, 8 isolates of S. saprophyticus, and 2 isolates of S. tests to provide results rapidly. Even the Vitek GPI cards, sm
capitiswerenegative.Anumberofauthorshavesuggestedthe whenrequiring.5hincubation,arenotrapidenough,because .
o
potential usefulness of including tests for lipases for the dif- resultsissuedbythelaboratorylateintheafternoonmaynot r
g
ferentiationofbacteriaingeneralandofstaphylococciinpar- reach the clinician or influence patient care till the following
/

ticular (17, 110, 201). Newly synthesized chromogenic sub- day. However, it is important to realize that commercial kits o
n
strates for the detection of esterases and lipases, including evolveandthatperformanceconcernsaredealtwithbymod-

A
propionate,decanoate,andlaurateestersof5-(4-hydroxy-3,5- ifications to formulae and/or software updates. It is therefore
p
dimethyoxyphenylmethylene)-2-thioxothiazoline-3-acetic acid importanttocheckwhichversionofthesystemisavailableand r
(SRA), available from Melford Laboratories, Chelsworth, Ip- howitsperformancerelatestopublisheddata. il
3
swich,UnitedKingdom,havebeenproposedforusewithbac- Thecompletiontimesforthedifferentidentificationschemes ,

terialisolates(161). varyfrom2hwiththeMicroScanRapidPosIDsystemto24 2
0
Stomatococcus. A description of the genus Stomatococcus, to72hwithsystemsthatrelyongrowth-dependenttests.The 1
whichcontainsonlyonespecies,Stomatococcusmucilaginosus, MicroScan Rapid Pos ID system requires a McFarland 0.5 9

isavailable(12,129,160,198,217).Strainsofthespeciesshow inoculumandreliesonthedetectionofenzymaticactivitywith b
y
themorphologicalcharacteristicsofthefamily:cocciarranged fluorogenic substrates and automated detection of fluores-
g
inpairs,tetrads,andclusters.TheyexhibitahighG1Ccontent cencebytheWalkAwayinstrument.RAPiDECStaph,relying u
of56to60mol%,neartothatofmembersofMicrococcus(64 onvisualobservationoffluorescencechange,canalsoprovide e
s
to75mol%)andunlikethatofstaphylococci(30to39mol%). identificationin2hbutrequiresaMcFarland4inoculumand t
Strainsofthespeciesshowalowcatalaseactivityandapositive canidentifyonlythreespecieswithoutadditionaltests.
benzidineactivity.Responsestometabolicandenzymatictests
resemblethoseofbothstaphylococciandmicrococci,e.g.,pro- StreptococcaceaeandRelatedOrganisms
duction of acid from a number of mono-, di-, and trisacchar-
ides(similartostaphylococci)andhydrolysisofmono-,di-,and The family Streptococcaceae and related organisms include
triamino acid conjugates (similar to Micrococcus). Enzyme theaerobicandfacultativelyanaerobicgram-positivecoccithat
tests used for the identification of S. mucilaginosus include generally give a negative result in the catalase test. These
hydrolysis of esculin (b-glucosidase), catalase, leucine-amino- organismsareencounteredinthemouthsandintestinaltracts
peptidases(LAP),andPYR.Identificationofmembersofthis ofhumansandanimalsandplayanimportantroleinthefood
taxon has been reported (12, 194, 195). Cross-infection be- industryasagentsofpreservationorspoilageoffermentation
tween patients has been studied by using phenotypic charac- products(220).Someofthetaxaarevirulentpathogens,caus-
ters,acidproductionfromcarbohydratesasdeterminedbyAPI ing pharyngitis, respiratory infections, skin and soft tissue in-
50CH,andMICs(227). fections, dental caries, infective endocarditis, and septicemia.
Identification schemes and kits. Reports on the identifica- Others play an important role in fermentation and preserva-
tion of staphylococci with commercial kits are summarized in tionprocessesofavarietyoffoodproducts.Isolatesarechar-
Table 7; when meaningful, identification of individual taxa is acterized by homo- or heterofermentative metabolism of car-
326 BASCOMB AND MANAFI CLIN.MICROBIOL.REV.
TABLE 7. ComparisonoftheaccuracyofidentificationsystemsfortheMicrococcaceae
%Correct
Identification Year Group No.ofisolates
(%correctwith Comments;(additionaltest);problematictaxa
system (reference) identified tested
additionaltests)
API20GPa 1994(181) CoNS 277 61.0 Bloodstreamisolates;S.epidermidis,otherCoNSspecies
APIID32Stapha 1995(108) CoNS 440 95.2 Clinicalisolates;S.capitis,S.hominis
APIID32Staph 1995(192) Staphylococci 89 82.1 Clinicalisolates
APIID32Staph 1995(192) Staphylococci 111 77.4 Culturecollectionisolates;CoNSspecies,including
S.hominis,S.schleiferi,S.warneri,S.xylosus
ATB32Stapha 1991(141) S.hyicus 54 83.3 Isolatesfrompigsandcattle
APIStapha 1982(87) CoNS 100 90.0 24–48hofincubation
APIStapha 1982(155) Micrococcaceae 300 30.0 Stockcultures,24–48hofincubation
APISTAPH-IDENTa 1983(5) CoNS 300 67.0(92.7) Clinicalisolates;(mostlynovobiocin)
APISTAPH-IDENT 1984(89) CoNS 120 79.2 Clinicalisolates,5-hincubation;S.haemolyticus,
S.hominis
APISTAPH-IDENT 1986(107) CoNS 175 88.0 S.epidermidis
APISTAPH-IDENT 1990(176) CoNS 55 100.0 Bloodisolates;reproducibilitystudy
D
APISTAPH-IDENT 1984(10) Micrococcaceae 414 (97.5) Clinicalisolates;(oxidase,susceptibilitytofurazolidone
o
andlysostaphin) w
APISTAPH-IDENT 1983(61) Staphylococci 188 80.9(90.4) Clinicalisolates;(24-hincubation) n
APISTAPH-IDENT 1983(143) Staphylococci 581 54.0 Bovineisolates;S.epidermidis,S.hominis lo
STAPH-IDENTa 1986(107) CoNS 175 88.0 Clinicalisolates;S.epidermidis a
d
STAPH-IDENT 1995(195) Micrococcaceae 1,106 81.1 StrainssubmittedtoCDCforidentification,stock
e
cultures,andacollectionofocularisolates d
S.aureus 162 77.2 f
r
S.cohnii 9 11.1 o
S.epidermidis 517 97.1 m
S.haemolyticus 61 75.8 h
S.hominis 57 82.5 t
t
Staph-TRAC(API) 1994(181) CoNS 277 73.0 Bloodstreamisolates;CoNSspeciesotherthan p
:
S.epidermidis //
Staph-TRAC(API)a 1990(157) Staphylococci 130 89.8 Bovineisolates cm
Staph-TRAC(DMS)a 1984(89) CoNS 120 88.3 Clinicalisolates,24-hincubation;S.haemolyticus,
r
S.warneri .a
Biologb 1993(162) Micrococcaceae 113 69.0–74.3 Stockisolatestestedattwoseparatelaboratories s
m
Gearyetal.scheme 1989(86) Staphylococci 559 94.6 Clinicalisolates,multipointinoculationonagarplates
MSGPc 1986(107) CoNS 175 86.4 Clinicalisolates;S.hominis,S.warneri,S.sciuri .o
MSGP 1994(97) CoNS 224 79.0 Clinicalisolates rg
S.epidermidis 44 95.0 /

S.haemolyticus 21 95.0 o
n
S.hominis 50 64.0
A
S.saprophyticus 11 100.0
p
S.warneri 39 79.0 r
MSGP 1991(130) Staphylococci 896 80.4(98.1) Culturecollection,clinicalandskinisolates;(includes il
3
probableidentificationswhenthecorrectspecieshas
,
notreachedrequiredlevelofidentification,buthas 2
beenincludedinthefourpossibletaxa) 0
1
S.aureus 58 91.0(100.0)
9
S.auricularis 22 91.0(100.0)
b
S.capitis 70 94.2(98.6) y
S.cohnii 49 82.4(100.0) g
S.epidermidis 152 82.0(98.0) u
S.haemolyticus 71 77.5(100.0) e
s
S.hyicus 22 82.0(96.0) t
S.intermedius 30 90.0(96.0)
S.kloosii 18 72.0(100.0)
S.lugdunensis 22 64.0(96.0)
S.saprophyticus 50 90.0(96.0)
S.warneri 55 63.6(90.9)
S.xylosus 32 66.0(94.0)
MSGP 1992(50) Staphylococci 370 73.4(92.0) Multicenterstudy;(requiringadditionaltests)
MSGP 1995(241) S.epidermidis 57 84.0(95.0) Bloodisolates
Non-S.epidermidis 16 62.0(70.0)
MSRapidGPc 1990(91) CoNS 342 88.3(96.2) Stockcultures
MSRapidGP 1994(97) CoNS 224 76.0 Clinicalisolates
S.epidermidis 44 91.0
S.haemolyticus 21 90.0
S.hominis 50 64.0
S.saprophyticus 11 100.0
S.warneri 39 77.0
Continuedonfollowingpage
VOL.11,1998 ENZYME TESTS FOR GRAM-POSITIVE COCCI 327
TABLE 7—Continued
%Correct
Identification Year Group No.ofisolates
(%correctwith Comments;(additionaltest);problematictaxa
system (reference) identified tested
additionaltests)
MSRapidGP 1991(130) Staphylococci 918 88.6(99.1) Culturecollection,clinicalandskinisolates;(includes
probableidentificationswhenthecorrectspecieshas
notreachedrequiredlevelofidentification,buthas
beenincludedinthefourpossibletaxa)
S.aureus 58 91.0(95.0)
S.auricularis 22 100.0(100.0)
S.capitis 70 88.6(100.0)
S.caprae 11 91.0(100.0)
S.cohnii 49 89.8(93.8)
S.chromogenes 19 68.0(94.0)
S.epidermidis 152 92.0(99.0)
S.haemolyticus 71 81.6(94.5)
S.hyicus 22 86.0(95.0) D
S.intermedius 30 87.0(90.0) o
S.kloosii 19 100(100.0) w
S.lugdunensis 24 62.0(70.0) n
S.saprophyticus 50 90.0(100.0) lo
a
S.warneri 55 83.6(92.7) d
S.xylosus 32 97.0(100.0) e
MSRapidGP 1992(50) Staphylococci 380 80.3(93.2) Multicenterstudy;(requiringadditionaltests) d

MSRapidGP 1992(221) Staphylococci 239 91.6(95.4) Stockculturesofclinicalisolates;(lowprobabilityof fr
o
identification) m
S.aureus 18 100.0(100.0)

S.capitis 21 85.7(90.5) h
t
S.cohnii 10 80.0(80.0) tp
S.epidermidis 40 95.0(100.0) :/
S.haemolyticus 20 100.0(100.0) /c
S.lugdunensis 14 92.9(100.0) m
S.saprophyticus 28 96.4(100.0) r.
a
S.warneri 25 96.0(96.0) s
S.xylosus 12 75.0(75.0) m
MSRapidGP 1995(108a) Staphylococci 69 82.6(91.4) Clinicalisolates;(lowprobability) .
o
MSRapidGP 1992(229) S.aureus 31 100.0 Clinicalisolates r
g
MSRapidGP 1995(182) S.aureus 20 90.0 Directidentificationofpositivebloodculture /

centrifugedpellets o
CoNS 95 53.7 n

MSRapidGP 1995(241) S.epidermidis 57 93.0(97.0) Bloodisolates A
OtherCoNS 16 38.0(49.0) p
r
RAPiDECStapha 1991(85) Staphylococci 124 60.5 Referencestrains il
RAPiDECStaph 1991(85) Staphylococci 121 88.4 Urineisolates 3
,
RAPiDECStaph 1991(165) Staphylococci Directdetectioninbloodcultures,modificationfrom 2
recommendedmethodbypredilutionofbloodbroth 0
mixturewithwater.CoNSspeciesdeterminationnot 1
9
pursued
b
CoNS 59 100.0 y
S.aureus 28 96.4 g
RAPiDECStaph 1994(112) Staphylococci 303 87.0 Clinicalandstockcultureisolates u
STAPHYtest 1991(211) Staphylococci 145 72.0 Stockcultures,24hincubation;manyCoNSspecies e
s
includingS.epidermidis,S.hominis,S.warneri, t
S.xylosus
Staph-Zymd 1992(191) CoNS 131 95.0 Clinicallysignificantisolates
Staph-Zym 1995(108) CoNS 440 97.5 Clinicalisolates;S.warneri
Staph-Zym 1991(239) Staphylococci 148 91.1 Bovineisolates;S.warneri,S.hominis
Staph-Zym 1995(192) Staphylococci 89 82.1 Clinicalisolates
Staph-Zym 1995(192) Staphylococci 111 99.0 Culturecollectionisolates;S.warneri
Staph-Zym 1989(139) S.hyicus, 85 100.0 Isolatesfrompigs,bovines,andcanines
S.intermedius
VitekGPI 1992(191) CoNS 131 95.0 Clinicallysignificantisolates
VitekGPI 1993(13) CoNS 500 77.0(86.0) Clinicalisolates;(goodconfidence-marginalseparation)
S.capitissubsp.captis 8 88.0(88.0)
S.epidermidis 322 89.0(92.0) S.capitissubsp.ureolyticusandS.lugdunensisarenotin
thedatabase
S.haemolyticus 68 71.0(95.0)
S.hominis 37 41.0(63.0)
S.saprophyticus 5 80.0(100.0)
Continuedonfollowingpage