GOING FOR THE GOLD: URINALYSIS AND BEYOND · GOING FOR THE GOLD: URINALYSIS AND BEYOND Joe Bartges, DVM, PhD, DACVIM, DACVN Professor of Medicine and Nutrition The University of Georgia

GOINGFORTHEGOLD:URINALYSISANDBEYONDJoeBartges,DVM,PhD,DACVIM,DACVNProfessorofMedicineandNutrition

[email protected]

• Partofaminimumdatabase• Evaluatewithurinarydisease:“Godmadeitgoldforareason”• Collectbyvoid,cystocentesis,catheterization

• Evaluatewithin30minutes• Ifnotpossible,refrigerate(2-8C)upto24hours• Re-warmtoroomtemperature

• Delaymayalterresults• Bacterialcontaminationordeath• Glucoseutilization• FalseincreaseinpHandprotein• Castsandcellsdeteriorate• Invivoprecipitationofcrystals

• Componentsofcompleteurinalysis• Urineappearance

• Color• Normalurineistransparentandyelloworamber• Intensityofcolorisrelatedtovolumeofurinecollectedandconcentrationofurine• Interpretincontextofurinespecificgravity(USG)• Diseasemayexistwithnormalcolor• Abnormalcolormaybecausedbypresenceofendogenousorexogenouspigments,butitdoesnot

providespecificinformation.Interpretationofsemi-quantitativereagentstrips,whicharecolorimetrictests,requiresknowledgeofurinecolorbecausediscoloredurinemayresultinafalsepositiveresult.Equineurinemayturnbrownafteraperiodoftime.

• Redurinedoesnotmeanblood(hematuria)• PositiveOCCULTBLOODonteststrip–canbeblood,hemoglobin,ormyoglobin• Pigment:e.g.myoglobin(clearserum)orhemoglobin(redserum)• Otherpigments:e.g.drugs

Table.PotentialcausesofdiscoloredurineURINECOLOR CAUSES URINECOLOR CAUSESYelloworamber Urochromes

UrobilinYellow-brownorgreen-brown

Bilepigments

Deepyellow HighlyconcentratedurineQuinacrine*Nitrofurantoin*Phenacetin*Riboflavin(largequantities)*Phenolsulfonphthalein(acidicurine)*

Browntoblack(brownorred-brownwhenviewedinbrightlightinthinlayer)

MelaninMethemoglobinMyoglobinBilepigmentsThymol*Phenoliccompounds*Nitrofurantoin*Nitrites*Naphthalene*Chlorinatedhydrocarbons*Anilinedyes*Homogentisicacid*

Blue MethyleneblueIndigocarmineandindigobluedye*Indicans*Pseudomonasinfection*Water-solublechlorophyll*Rhubarb*Toluidineblue*Triamterene*Amitriptyline*Anthraquinone*Bluefooddye*

Colorless Verydiluteurine(diuretics,diabetesmellitus,diabetesinsipidus,glucocorticoidexcess,fluidtherapy,overhydration)

Green MethyleneblueDithiazanineUratecrystalluriaIndigoblue*Evan’sblue*BilirubinBiliverdinRiboflavin*Thymol*Phenol*Triamterene*Amitriptyline*Anthraquinone*Greenfooddye*

Milkywhite LipidPyuriaCrystals

Red,pink,red-brown,red-orange,ororange

HematuriaHemoglobinuriaMyoglobinuriaPorphyrinuriaCongoredPhenolsulfonphthaleinNeoprontosilWarfarin(orange)*Foodpigments(rhubarb,beets,blackberries)*

Carbontetrachloride*

Brown MethemoglobinMelaninSulfasalazine*Nitrofurantoin*Phenacetin*Naphthalene*Sulfonamides*Bismuth*Mercury*Feces(rectal-urinaryfistula)Favabeans*

PhenazopyridinePhenothiazine*Diphenylhydantoin*BromsulphaleinChronicheavymetalpoisoning*Rifampin*Emodin*Phenindione*Eosin*Rifabutin*Acetazolamide*Redfooddye*

Rhubarb*Sorbitol*Metronidazole*Methocarbamol*Anthracincathartics*Clofazimine*Primaquine*Chloroquine*Furazolidone*Coppertoxicity

Orange-yellow HighlyconcentratedurineExcessurobilinBilirubinPhenazopyridineSulfasalazine*Fluoresceinsodium*Flutamide*Quinacrine*Phenacetin*2,4-d*Acetazolamide*Orangefooddye*

*Onlyobservedinhumanbeings• Turbidity

• Urineistypicallyclear• Maybelesstransparentwithpigmenturia,crystalluria,hematuria,pyuria,lipiduria,orother

compoundssuchasmucous.• Theincreasedturbiditymaydisappearwithcentrifugationofthesampledependingonthe

causeofincreasedturbidity.• Odor

• Normalurinehasaslightodorofammonia;however,theodorisdependentonurineconcentration.

• Somespecies,suchascats(felinine)andgoats,havepungenturineodorbecauseofurinecomposition.

• Bacterialinfectionmayresultinaverystrongodorduetopyuriaandastrongammoniaodorifthebacterialorganismproducesurease.

• Centrifugefor3-5minutesat1500-2000rpm

• Pouroffsupernatant• CandoUSG• Usefordipstick(semi-quantitativechemicaltesting)

• Leavesmallpelletwithlittleurineintubeformicroscopicexamination• Dipstick(semi-quantitative,colorimetricreagentstrips)

• ReagentstripssuchasMultistix®orChemstrip®canbeusedtoperformseveralsemi-quantitativechemicalevaluationssimultaneously.

• DetermineurinepH,proteinglucose,ketones,bilirubin/urobilinogen,andoccultblood.• Somereagentstripsincludetestpadsforleukocyteesterase(fordetectionofwhiteblood

cells,(WBC)),nitrite(fordetectionofbacteria),andUSG;thesearenotvalidinanimalsandshouldnotbeused.

• Reagentstripsareadverselyaffectedbymoistureandhavealimitedshelflife.Bottlesshouldbekepttightlycapped,andunusedstripsshouldbediscardedaftertheirexpirationdate.

• pH• Variesfrom5.0-9.0• Accuratetowithin0.5pHunits

• Areadingof6.5meanstheactualpHislikelytobebetween6.0and7.0• Typicallyacidicindogsandcatsandalkalineinhorsesandruminants,

• Variabledependingondiet,medications,orpresenceofdisease.• Falselyincreasedifleftatroomtemperature• Abacterialurinarytractinfectionwithaurease-producingmicrobewillresultin

alkaluria.• UrinepHwillaffectcrystalluriabecausesomecrystals,suchasstruvite,formin

alkalineurine,whileothercrystals,suchascystine,forminacidicurine.• Protein

• Methods• Dipstick

• Detectsprimarilyalbumin• 30-3,000mg/dl

• Falsenegativeforotherproteins(e.g.globulins)• Falsepositivewithalkaluria

• Sulfasalicylicacidprecipitationtest• Detectsalbuminandglobulins• Usedasverification• Notaccurate

• Quantitative• Measurealbuminorproteinina24-hoururinesample• Urineproteintourinecreatinine(UPC)

• <0.2=normal• 0.2–0.4(cats)and0.5(dogs)=borderline• >0.4(cats)and>0.5(dogs)=abnormal

• Microalbumin• Semi-quantitativeELISAtestformicroalbuminuria

• 1-30mg/dl• Differentkitsfordogsandcats

• Interpretation

• Smallamountisnormal• Alkaluriacangiveslightfalsepositivereaction• Canbeseenwithinflammation,hemorrhage,orglomerulardisease.• ApositivereactionmustbeinterpretedinlightofUSG,pH,andurinesediment

examination.• Forexample,atraceamountofproteininconcentratedurineislesssignificant

thanatraceamountofproteinindiluteurine.• Presenceofotherproteins,suchasBence-Jonesproteins,willgivefalse

negativeresults.• Glucose

• Notpresentnormally• Renalthresholdis>180mg/dlinmostspecies;>240mg/dlincats.• Witheuglycemia,theamountoffilteredglucoseislessthantherenalthresholdandall

ofthefilteredglucoseisreabsorbedintheproximalrenaltubules.• Glucosuriacanresultfromhyperglycemia(duetodiabetesmellitus,excessive

endogenousorexogenousglucocorticoids,orstress)orfromaproximalrenaltubulardefect(suchasprimaryrenalglucosuriaorFanconisyndrome).

• Ifglucosuriaispresent,bloodglucoseconcentrationshouldbedetermined.• Ketones

• Detectsacetateandacetoacetate,butnotbeta-hydroxybutyrate.• Associatedwitheitherprimaryketosis(ruminants),ketosissecondarytodiabetes

mellitus(smallanimals),andoccasionallywithprolongedfastingorstarvation.• Afalsepositivereactioncanoccurwithpresenceofreducingsubstancesinurine.

• Bilirubin/urobilinogen• Bilirubinuriaoccurswhenconjugatedbilirubinexceedsrenalthresholdaswithliver

diseaseorhemolysis.• Indogswithconcentratedurine,asmallamountofbilirubincanbenormal.• BilirubinuriaisALWAYSabnormalincats• Atablettest,Ictotest®,ismoresensitive• Pigmenturiamayresultinafalsepositivereaction.

• Urobilinogen,formedfrombilirubinbyintestinalmicroflora,isabsorbedintotheportalcirculationandisexcretedrenally• Notspecificenoughtobeclinicallyuseful.

• Occultblood• A“pseudoperoxidase”methodtodetectintactredbloodcells(RBC),hemoglobin,and

myoglobin.• Apositivereactioncanbeduetohemorrhage(hematuria),intravascularhemolysis

(hemoglobinuria),ormyoglobinuria.• Thelattertwoprocessescanbedistinguishedbyexaminationofplasma;plasma

willappearpinktoredafterintravascularhemolysis,whilemyoglobinisrapidlyclearedfromplasmaresultinginclearplasma.

• Aswithothercolorimetrictestpads,discoloredurinemayyieldfalsepositiveresults.

• Apositiveresultshouldbeinterpretedwithmicroscopicexaminationofurinesediment.

• Othersemi-quantitativetestpads• Nitrite,leukocyte,andurinespecificgravity–therearetestpadsfordetectionofnitrite

(frombacterialinfection),leukocytes(leukocyteesterasefromwhitebloodcellspresentinurine),andurinespecificgravity.Thesearenotreliableforuseinanimals.

• Urinespecificgravity• Indirectmeasureofosmolality,whichisabettermeasureofconcentration

• Doonwholeurineorsupernatantaftercentrifugationifurineisdiscolored• Determinedusingarefractometerdesignedforveterinarysamples,whichincludesascalecalibrated

specificallyforcaturine.• USGforspeciesotherthancatsshouldbedeterminedusingthescalefordogs• Interfaceofdarkandlight• Massrelativetothemassofdeionizedwater(USG=1.000)

• Highlyvariable,dependingonfluidandelectrolytebalanceofthebody• Interpretationdependsonclinicalpresentationandserumchemistryfindings.• Ananimalthatisdehydratedorhasothercausesofprerenalazotemiawillhavehypersthenuric

urinewithaUSG>1.025-1.040(dependingonspecies).• Diluteurineinadehydratedorazotemicanimalisabnormalandcouldbecausedbyrenalfailure,

hypoadrenocorticism,hyperadrenocorticism,hypercalcemia,diabetesmellitus,hyperthyroidism,diuretictherapy,ordiabetesinsipidus.

• GlucosuriaincreasestherefractiveindexofurineresultinginanincreasedUSGdespiteincreasedurinevolume.

• UrineSediment

• Followingcentrifugation,decantsupernatantleavingapproximately0.5mlofurineandsedimentinthetipoftheconicaltube.

• Re-suspendpelletbytappingthetipoftheconicaltubeagainstthetableseveraltimes.• Transferafewdropsofthesedimenttoaglassslide,andacoverslipisapplied.• Examinationofunstainedurineisrecommendedforroutinesamples.• Microscopicexaminationisperformedat100X(forcrystals,casts,andcells)and400X(forcells

andbacteria)magnifications.• Contrastofthesampleisenhancedbyclosingtheirisdiaphragmandloweringthecondenser

ofthemicroscope.• StainssuchasSedistain®andnewmethylenebluecanbeusedtoaidincellidentificationbut

tendtodilutethespecimenandintroduceartifactssuchasstainprecipitateandcrystals.• Interpretresultswithurinedipstickevaluation,USG,andurinespecimenhandling• Cells

• Redbloodcells• Smallandroundandhaveaslightorangetintandsmoothappearance.• Normalurineshouldcontain<5RBC/fieldat400Xmagnification.• IncreasedRBCinurine(hematuria)indicateshemorrhagesomewhereintheurogenital

system;however,samplecollectionbycystocentesisorcatheterizationmayinducehemorrhage.

§ Whitebloodcells• SlightlylargerthanRBCandhavegrainycytoplasm.• Normalurineshouldcontain<5WBC/fieldat400Xmagnification.• IncreasedWBC(pyuria)canoccurduetoinflammation,infection,trauma,orneoplasia.• CatheterizationorcollectionofvoidedurinemayintroduceafewWBCfromthe

urogenitaltract.

Whitebloodcell(left)and2redbloodcells(right)\

• Epithelialcells

• Transitionalepithelialcells,acommonurinecontaminantderivedfromthebladderandproximalurethra,resembleWBCbutarelarger.• Theyhaveagreateramountofgrainycytoplasmandaround,centrallylocated

nucleus.• Inavoidedurinesample,squamousepithelialcellsmaybeobserved.

• Theyarelarge,ovaltocuboidalinshape,andmayormaynotcontainanucleus.• Occasionally,neoplastictransitionalcellsmaybeobservedinananimalwitha

transitionalcellcarcinomaorneoplasticsquamouscellsmaybeobservedinananimalwithasquamouscellcarcinoma.

• Cylindruria(casts)• Elongated,cylindricalstructuresformedbymucoproteincongealingwithinrenaltubules

andmayormaynotcontaincells.• Hyalinecastshaveparallelsidesandroundedends,andarecomposedofmucoprotein.

• Theymayoccurwithfever,exercise,andrenaldisease.• Cellularcasts

• Epithelialcellularcastsformfromentrapmentofsloughedtubularepithelialcellsinthemucoprotein;theymaybeobservedwithrenaldisease.

• Granularcastsarethoughttorepresentdegeneratedepithelialcellularcasts.• Mostcommontypeofcast

• Waxycastshaveagranularappearance,andarethoughttoarisefromlong-standinggranularcasts.• Theytypicallyhavesharpborderswithbrokenends.

• Afewhyalineorgranularcastsareconsiderednormal.However,presenceofcellularcastsorothercastsinhighnumbersindicaterenaldamage,andmaybeoneoftheearliestlaboratoryabnormalitiesnotedwithtoxicdamagetorenalepithelialcells(eg,gentamicin,amphotericinB).• Erythrocytecastsformbecauseofrenalhemorrhage.• WBCcastsoccurbecauseofrenalinflammation,aswithpyelonephritis.

• Fattycastsarenotcommon,butcanbeobservedwithdisordersoflipidmetabolism,suchasdiabetesmellitus.

• Infectiousorganisms

• Presenceofbacteriainurinecollectedbycystocentesisindicatesinfection.• Smallnumbersofbacteriafromthelowerurogenitaltractmaycontaminatevoided

samplesorsamplescollectedbycatheterization,anddonotindicateinfection.• Bacterialrodsaremosteasilyidentifiedinurinesediment.• Particlesofdebrismaybemistakenforbacteria.• SuspectedbacteriacanbeconfirmedbystainingurinesedimentwithGram’sstain;

however,aerobiccultureisbesttoconfirmabacterialurinarytractinfection.• Rarely,yeastandfungalhyphaeandparasiticovamaybeobservedinurinesediment.

• Theirpresenceisnotalwaysassociatedwithclinicaldisease.• ParasiticovaobservedincludeStephanusdentatus,Capillariaplica,Capillariafelis,

andDioctophymarenale.• Additionally,microfilariaeofDirofilariaimmitismaybeobservedinurinesediment.

WBCwithbacteria Capillariafelis

• Crystals

• Manyurinesedimentscontaincrystals.• ThetypeofcrystalpresentdependsonurinepH,concentrationofcrystallogenic

materials,urinetemperature,andlengthoftimebetweenurinecollectionandexamination.

• Crystalluriaisnotsynonymouswithurolithiasis,andisnotnecessarilypathologic.• Furthermore,urolithsmayformwithoutobservedcrystalluria.• Struvite

• Struvitecrystalsappeartypicallyas“coffin-lids”or“prisms”;however,theymaybeamorphousinappearance.• OccurinalkalinepH

• Struvitecrystalsarecommonlyobservedincanineandfelineurine.• Herbivoresoftenhavestruvitecrystalluriaduetoalkaluria

• Struvitecrystalluriaindogsisnotaproblemunlessthereisaconcurrentbacterialurinarytractinfectionwithaurease-producingmicrobe.• Withoutaninfection,struvitecrystalsindogswillnotbeassociatedwith

struviteurolithformation.• Someanimals,suchascats,however,doformstruviteurolithswithoutabacterial

urinarytractinfection.• Intheseanimals,struvitecrystalluriamaybepathologic.

• Calciumoxalate• OccursinacidicpH• Twoforms:

• Monohydrate:“dumb-bell”inshape.• Dihydrate:“envelope”or“squarewithX”

• Occurslesscommonlyindogsandcats• Anunusualformofcalciumoxalatecrystalsistypicallyseeninassociationwith

ethyleneglycoltoxicity.• Maybeassociatedcalciumoxalateurolithformation.

• Commoninhorsesandcattle• Calciumcarbonate

• Commoninhorses,rabbits,guineapigs,andgoats• OccursinalkalinepH

• Variablysizedandoftenappearaslargespheroidswithradialstriations• Mayalsobesmallercrystalswithround,ovoid,ordumbbellshapes

• Colorlesstoyellow-brown• Ammoniumurate

• OccurinacidicpH• Yellow-brownsphereswithirregular,spinyprojections;however,theymayalsobe

amorphous• Ammoniumaciduratecrystalssuggestliverdisease(eg,portosystemicshunt).• Certainspecies,suchasbirdsandreptiles,andcertainbreedsofdogs,specifically

Dalmatians,cannormallyhaveammoniumaciduratecrystalluria.• Cystine

• Cystinecrystalsaresix-sidedandofvariablesize.• Theyoccurinacidicurine.• Presenceofcystinecrystalsrepresentsaproximaltubulardefectinaminoacid

reabsorption.• Cystinuriahasbeenreportedtooccurinmanybreedsofdogsandrarelyincats,but

Dachshunds,Newfoundlands,Englishbulldogs,andScottishterriershaveahighincidenceofcystineurolithiasis.

• Bilirubin• Bilirubincrystalsoccurwithbilirubinuria;however,thesemaybenormalinsmall

numbersindogs.

Ammoniumuratecrystals Bilirubin Cystine

Calciumoxalatemonohydrate Calciumoxalatedihydrate Struvite

• Lipid• Fatdropletsarecommonlypresentinurinefromdogsandcatsandmaybemistakenfor

RBC.However,theyoftenvaryinsizeandtendtofloatonadifferentplaneoffocusthantheremainderofthesediment.Theyarenotconsideredtobepathologic.

• Spermatozoa• Spermatozoamaybeobservednormallyinurinecollectedfrommaledogs.

• Artifact• Occasionally,plantmaterialmaybeobservedinurinesamplescollectedbyvoiding.

Whenpresent,theyindicatecontaminationoftheurinesample,andarenotpathologic.

• Talcpowdergranulesmaybeobservedinvoidedsamplesifthecollectoriswearinggloves

• Enzymuria.• Enzymaticactivity,usedasamarkerforAKI,belongtoenzymesthatarefoundwithintherenaltubularcells.• Theseenzymesaretoolargetobefilteredthroughanormalglomerulus,andsointheabsenceofprofound

glomerulardisease,ariseintheurinaryactivityofsuchenzymesistypicallycausedbyacutedamagetothetubulesandleakagefromthetubularcells.

• Urinaryenzymes,GGT,andNAGarethemostcommonlyusedandmostpracticalenzymestoassessurinaryactivity.• NAGisfoundwithintheproximaltubularlysosomesandGGTwiththeproximaltubulebrushborder.• Theactivityoftheseenzymesisasensitivemethodofdetectingacutetubularkidneyinjury,moresensitive

thanchangesinglomerularfiltrationrate,serumbiochemistry(azotemia)andclinicalsigns.• Changesinurinaryenzymeconcentrations,GGTandNAG,canbeestimatedbyenzymetocreatinineratioson

spoturinesamples,deeming24-hoururinecollectionsnotabsolutelynecessary.• Examplesofsuchcasesincludetheuseofrenaltoxicchemotherapeuticagents,theuseofaminoglycosides,a

veryrecentoverdoseofanon-steroidalanti-inflammatorydrug(NSAID)ortheuseofanNSAIDinarenalcompromisedpatient.

• Point-of-caretestingforurinarytractinfections• Thesetestsmaydetectpresenceofbacteria,determinetypeofbacteria(Gram+versusGram-),identifythe

bacterialorganism,and/orprovidesusceptibilitytoalimitednumberofantimicrobialagents• Otherinfectiousagents

• UrinemaybesubmittedforLeptospirosisPCRfordiagnosisofleptospirosisorforfungalPCRfordiagnosisoffungalinfectionssuchasBlastomycosis,Aspergillus,andHistoplasmosis

• Bladdertumorantigentest(VBTA)• TheVBTAcanbeusedasascreeningtestfortransitionalcellcarcinomaindogs.• Theresultsarenotspecificandnon-neoplasticdisease(e.g.urinarytractinfections,hematuria,etc)cangive

positiveresults.• Anegativetest;however,ismeaningfulinthatatransitionalcellcarcinomaisnotlikelytobepresent.• Thistestmaybeusefulforroutinescreeningofdogsathigherriskofdevelopingtransitionalcellcarcinoma(e.g.

Scottishterriers)thatdonothaveothersignsorlaboratoryfindingsoflowerurinarytractdisease.