Urine Analysis Presentation

Urine Examination&

Analysis

Assigned by: Dr.Javeria Khan

Presented by: Dr.Noor-ul-Ain Sarwar

Contents

1. Collection of sample and preservation.

2. Gross Examination.

3. Determination of specific gravity.

4. Biochemical Analysis.

5. Microscopic Examination.

6. Disease Interpretations

Introduction

• Urinalysis a very useful tool to evaluate healthy and diseased animals. It provides valuable information about the urinary system. There are so many examples of diseases in which specific urine picture can be seen. for example:

A. Kidney diseases: Abnormal specific gravity Proteinurea. Cast Leukocyte Erythrocytes

B. Bladder infection: Proteinuria Leukocytes Bacteria

C. Neoplasia: Exfoliated neoplastic cells Hematuria

D. Liver diseases: Billirubinuria Altered urobilinogen Bilirubin crystals

E. Hemolysis: Post parturient hemoglobinuria Bovine bacillary hemoglobinuria Anthrax Increased urobilinogen

F. Diabetes mellitus: Glycosuria Increased volume, increased specific gravity Ketonurea

G. Diabetes insipidus: Decreased specific gravity

H. AcidosisI. Alkalosis

Collection of Urine

1. Collection of urine sampleCollection can be done either by clean catch

method,catheterization or through cystocentesis.

Sample size: 15- 20mL

Best time for analysis:

A fresh urine sample is preferred for analysis.Ideally urinalysis should be performed within 30 min of sample collection.If delay of examination then following changes takes place:Urea is converted into ammonia that makes the

sample alkaline.Formed elements(cells,casts)are dissolved.

Precautions while collection of urine: Morning samples are most likely to contain

constituents of diagnostic significance.Fluid consumption during the day dilutes the urine

resulting in decreased specific gravity.Collect mid stream urine.In case of diabetes mellitus, sample should be

collected 2 hours after feeding and fasting.For nephritis,use only morning samples.Direct collection is the preferable method in large

animals.although catheterization and cystocentesis

provide high quality of uncontaminated sample but are associated with tissue trauma of varying degree.

Preservation: Store samples in refrigerator at 8 degree C.(warm

at room sample before analysis). Precautions regarding refrigeration:

Maximum upto 12 hoursIt slightly increases specific gravity.It also interferes with tests using enzymes for

reaction.

Chemical preservationCertain chemicals are used with limitations.

a. Toluene:

Quantity: 2ml/100ml of urine for 24 hours.

Only cover urine surface, don't dissolve in urine.

Limitation :Interferes with ketone bodies determination.

b. Thymol:

Quantity: a small lump can preserve for several days.

Limitation: it gives false positive protein reaction.

c. Formaline:

Quantity: 1drop of 40% formalin for 30ml urine for 24 hours.

Limitation: It interferes with glucose reaction.

c. Metaphosphoric acid:

When ascorbic acid is to be determined from the urine. It is added with a ratio of 1:5 ( 1 part 10% aqueous solution of metaphosphoric acid and 5 parts of urine sample.

Collection

2. Gross Examination

a) Volume:

species

Normal values

in liters

Horse 4.7

Cattle 14.2

Sheep, Goat and Dog

Interpretation

i. Increased volume-Polyurea:

Physiological: Increased water consumption Diuretics Parenteral fluid therapy

Pathological: Chronic progressive renal failure Diabetes mellitus Diabetes insipidus Chronic pylonephritis

Pyometra

ii. Decreased volume-oligouria:Physiological:

Less water intake. High environmental temperature. Panting Dehydration

Pathological: Acute renal disease Urolithiases fever Shock Severe nephritis Edema

b. Color:

Colour of the urine is due to the concentration

of urochromes.Always consider color in association of volume and specific gravity of urine.

Normal Colour:

Freshly voided urine is clear and may range in color from light pale yellow to amber(gold)or straw Colour except horses which have turbid color urine due the presence of calcium carbonate crystals and mucin.

InterpretationVarious color could be:

i. Less to pale yellow: End stage renal disease Increased uptake of water Diabetes insipidus Hyperadrenocorticism

ii. Dark yellow to yellow brown: Acute nephritis Dehydration Bilirubin

iii. Yellowish brown; In birds ,yellowish green urates indicates

hemolysis or liver disease. Bilirubin

iv. Red: Hematuria Hemoglobinuria

v. Brown to brownish black: Hemoglobin(hemoglobinuria,post parturient

hemoglobinuria,bacillary hemoglobinuria) Myoglobin(Monday morning disease) Melanin

vi. Green: Biliverdin Phenol poisoning

c. Odor Interpretation Ammonia-like :Ammonia-like : Urea-splitting bacteriaUrea-splitting bacteriaFoul, offensive :Foul, offensive : Old specimen, pus or Old specimen, pus or

inflammation inflammation Sweet : Sweet : GlucoseGlucose Fruity : Fruity : KetonesKetones Maple syrup-like: Maple syrup-like: Maple Syrup Urine DiseaseMaple Syrup Urine Disease

d.d. Color:Color:

Colorless Colorless Diluted urineDiluted urine Deep YellowDeep Yellow Conc. Urine, Riboflavin.Conc. Urine, Riboflavin. Yellow-GreenYellow-Green Bilirubin / BiliverdinBilirubin / Biliverdin RedRed Blood / HemoglobinBlood / Hemoglobin Brownish-redBrownish-red Acidified Blood (Actute Acidified Blood (Actute

GN)GN) Brownish-blackBrownish-black Homogentisic acid Homogentisic acid

(Melanin)(Melanin)

3. Specific Gravity

The ability of kidneys to concentrate the urine.Determination: It can be determined by the use of refractometer

or urinometer.the steps are as under: Temperature of the urine must be 20-25°C. cylinder used for floatation of urinometer

should be large enough in diameter so that urinometer can flow in it.

Place the urinometer in cylinder containing the urine .Rotate it to prevent its touching to the sides.

Read the scale on the bottom of urinometer

and record it in decimals.

Urinometer

Refractometer

Species Specific Gravity

Horse 1.020 – 1.050

Cattle 1.025 – 1.045

Sheep & Goat 1.015 – 1.024

Dog 1.015 – 1.045

Birds 1.005 – 1.020

Interpretation

i. Increased specific gravity: Acute interstitial nephritis Cystitis Liver failure Diabetes mellitus Glomerulonephritis

ii. Decreased specific gravity: Chronic interstitial nephritis Diabetes insipidus Pylonephritis uremia

Chemical AnalysisChemical Analysis

3. Biochemical Analysis

For biochemical analysis urine must be uncentrifuged.

a. pH:

i. Acidic pH Interpretation: Normal in carnivores. Nursing calves & foals. Excessive diet in protein. Hypokalemia.

ii. Alkaline pH Interpretation:

Normal in herbivores Stale urine sample becomes alkaline. Cystitis.

Normal pH

Species pH

Horses 8

Cattle 7.4-8.4

Sheep & Goat 7-8.2

Dog & Cat 5.5-7

Birds 6-8

b. Protein Determination:

For Protein determination:

a) Reagent strips(dip sticks)

b) Acid prepitation Tests:

i. Nitric Acid Precipitation Test

OR Robert’s Tests:

Principle:

Precipitation of protein occur by concentrated acid

Procedure:

Take 2ml of Robert’s reagent in a test tube.Place 2ml of urine.Wait for few minutes.

Result:

A positive test is indicated by a white ring at the zone of contact of 2 fluids.

Interpretation of Protein determination

Hemoglobinuria.Myoglobinnuria.Pylonephritis.Cystitis UrolithiasesInflammation,Hemorrhage, Glomerular disease.

c. Glycosuria determination

Now a days strips and glucometers are available. Chemical method is Benedict’s test.

Benedict’s test:

Principle:

It depends upon the reducing sugars present in the urine to react with copper sulphate to reduce cupric ions to cuprous oxide giving color.

Composition of Benedict’s Reagent

Copper sulphate 17.3 g

Sodium citrate 173 g

Sodium carbonate 100 g

Distilled water

(To make volume)

1000 mL

Procedure:

Take 5mL of Benedict’s reagent in a test tube.Add 8 drops of urine to the reagent.Mix the 2 fluids.Heat it with constant shaking till boiling.• Result:

Positive Blue color

Negative Orange to brick red or brown

Interpretation

HyperglycemiaAfter general anesthesiaChronic liver diseasesEnterotoxaemia in sheep.

d. Ketonurea Determination

Ross test:Principle:It is based on that the sodium nitroprusside is

decomposed to:Sodium ferrocyanideSodium nitrateFerric hydroxideResults:Purple coloration

Procedure:

Place half inch layer of powdered reagent in test tube.

Add 5mL of urine.Agitate the 2 components in the test tube.’Overlay 1-2 ml of ammonium hydroxide over

the mixture.Wait for 4-5 minutes.Development of purple color indicates the

presence of ketone bodies in urine.

Interpretations

Diabetes mellitusHigh fat dietStarvationImpaired liver functionsAfter ether chloroform anesthesiaMilk fever

e. Hematuria detection

Benzidine test:Take 2mL of glacial acetic acid in a test tube.Add small amount of Benzidine reagent.Add 1 ml of urineAdd 1 mL of fresh hydrogen per oxide.Wait for 5 minutes.Result:• Green or blue color development.

Interpretation• Acute nephritis

• Urolithiases

• Cystitis

• Tumor of the urethra

• Severe infections like, anthrax, leptospirosis,infectious canine hepatitis.

• Chemicals like copper, mercury or phenol poisoning.

• Parasites like Dicroflaria immitus,Dictophyma renale,Capillaria plica.

f. Billirubinuria determination

Foam test:

Procedure:Take 1-2 mL of urine in a test tube.Shake it vigorously.

Result:

• Appearance of yellow, greenish yellow or brown colour foam above the surface of urine indicates presence of Bilirubin.

Interpretation:

Infectious canine hepatitis,leptospirosis.NeoplasiaObstruction of bile duct.Jaundice

Calcium Determination

Sulkowitch test:

Calcium present in urine reacts with sulkowitch reagent ,ppt in the form of calcium oxalate.

Procedure:Take 5mL distilled water & add 5mL urine in

1 test tube as control.In another test tube,mix equal amount of urine

& sulkowitch reagent.

Result:Compare the 2 test tubes in light after 2-10 min.

Interpretation:Increased:After Ca administration.HyperthyroidismHypervitaminosis

Decreased: In bovines, it is not reliable. In canines,pre-renal tetany.hypothyroidism

5) Microscopic examination

Purpose:Recognition of cells for urinary tract

infections.Exfoliative cytology of tumors.

Procedure; Centrifuge sample @ 1500 rpm for 2-3 min. Pour off the supernatant. Place a drop of sediment on slide and cover it

with a cover slip. Observe the slide @ 10x and 40x.Result variations:i. Voided sample: more cellular, bacterial

contamination.ii. Catheterized sample: increased transitional cell

content, iatrogenic hemorrhage.iii. Cystocentesis: least extraneous contamination,

more specific for changes in the tract,

Interpretations:• Epithelial cells in neoplasia diagnosis.

• more than 5RBCs/HPF indicate Hematuria.

• Leukocytes indicate infection(pyouria).

• More than 5/HPF.

• Elongated structures like casts indicate presence of Urolithiases.

• 10,000 bacterial rods/ml and >100,000 bacterial cocci/mL of urine are required to consistently find bacteria in a urine sample using light microscopy. and readings are normally below this.

Some of the drugs that excreted in urine also appear in crystals.e.g:

Sulfonamide crystals spherical with spikes.Ampicillin crystals form long needle lik

arrays.Calcium oxalate crystals are like colorless

squares indicate: UrolithiasesEthylene glycol toxicosis

Cytological ExaminationCytological Examination

• Staining:Staining:

– PapanicolauPapanicolau– Wright’sWright’s– ImmunoperoxidaseImmunoperoxidase– ImmunofluorescenceImmunofluorescence

Staining:

WRIGHT STAIN PROCEDURE:Make a air dried smear.Fix it in methanol for 30 sec.Take a disposable pipette and flood the Wright Stain

on the appropriately labeled slides.Wait for 3 min. Place 1ml oxidizing Wright Stain and Wright Stain

Buffer Mixture on Wright stained slides laying on slide rack (Displacing the Wright Stain off the slides with the pipette filled with Wright Stain/buffer mixture and viewing a metallic sheen on the top of slides.)____

Staining:

Wait for 6 min. Place slides in Wright Stain Buffer for 1.5

minutes.wait for 1.5 minutes. Rinse, dry and examine under oil immersion

lens,100x.

Parasites

Capillaria plica

Dioctophyme renale.

Trichuris

RBCs Cast - HistologyRBCs Cast - Histology

RBCs CastRBCs Cast

WBCs CastWBCs Cast

Tubular Epith. CastTubular Epith. Cast

Granular CastGranular Cast

Hyaline CastHyaline Cast

Waxy CastWaxy Cast

Fatty CastFatty Cast

Crystals

Calcium Oxalate CrystalsCalcium Oxalate Crystals

Triple Phosphate CrystalsTriple Phosphate Crystals

Urate CrystalsUrate Crystals

Leucine CrystalsLeucine Crystals

Cystine CrystalsCystine Crystals

Bilirubin

Ammonium Biurate CrystalsAmmonium Biurate Crystals

Cholesterol CrystalsCholesterol Crystals

Cytology

carcinoma

Cytology: Polyoma (Decoy Cell)Cytology: Polyoma (Decoy Cell)

Cytology: Squamous Cell Ca.Cytology: Squamous Cell Ca.

Cytology: Renal Cell Ca.Cytology: Renal Cell Ca.

Cytology: Prostatic CarcinomaCytology: Prostatic Carcinoma

Cytology

Cytology: NormalCytology: Normal

Cytology: ReactiveCytology: Reactive

Tubular Epithelial CellsTubular Epithelial Cells

Hematuria

Transitional CellsTransitional Cells

Oval Fat BodyOval Fat Body

Transitional CellsTransitional Cells

LE CellLE Cell

Squamous cell

CytomegalovirusCytomegalovirus

YeastsYeasts

BacteriaBacteria

Amorphous Substance

Bacilli

Mucous

Interpretations of Urine Analysis

ProteinuriaCasts & cellsHematuriaHemoglobinuriaMyoglobinuriaPyuriaBacteriuriaCrystalluriaGlycosuriaKetonuriaParasites

6.Interpretation Of

Diseases of

Urinary System

Common Findings in:Common Findings in:

Acute Tubular NecrosisAcute Tubular NecrosisMicroscopic:Microscopic:Renal tubularRenal tubular epithelial cellsepithelial cellsPathological casts.Pathological casts.

Microscopic:Microscopic:Renal tubularRenal tubular epithelial cellsepithelial cellsPathological casts.Pathological casts.

GlucoseGlucose

BilirubinBilirubin

KetonesKetones

S.G.S.G.

BloodBlood

ProteinProtein

UrobilinogenUrobilinogen

DecreasedDecreased

+ / -+ / -

Common Findings inCommon Findings in::

Acute GlomerulonephritisAcute Glomerulonephritis

Microscopic:Microscopic:Erythrocytes (dysmorphic)Erythrocytes (dysmorphic)Erythrocyte castsErythrocyte castsMixed cellular castsMixed cellular casts

GlucoseGlucoseGlucoseGlucose

BilirubinBilirubinBilirubinBilirubin

KetonesKetonesKetonesKetones

Specific GravitySpecific GravitySpecific GravitySpecific Gravity

BloodBloodBloodBlood

pHpHpHpH

ProteinProteinProteinProtein

UrobilinogenUrobilinogenUrobilinogenUrobilinogen

NitriteNitriteNitriteNitrite

Leukocyte EsteraseLeukocyte EsteraseLeukocyte EsteraseLeukocyte Esterase

IncreasedIncreased

Common Findings inCommon Findings in::Chronic GlomerulonephritisChronic GlomerulonephritisMicroscopic:Microscopic:Pathological castsPathological casts

(broad waxy casts, RBCs)(broad waxy casts, RBCs)

Microscopic:Microscopic:Pathological castsPathological casts

(broad waxy casts, RBCs)(broad waxy casts, RBCs)

pHpHpHpH

DecreasedDecreased

IncreasedIncreased

Acute PyelonephritisAcute Pyelonephritis

Microscopic:Microscopic:BacteriaBacteriaLeukocytesLeukocytesLeukocyte, granular, andLeukocyte, granular, and

waxy castswaxy castsRenal tubular epithelialRenal tubular epithelial

cell castscell casts

Microscopic:Microscopic:BacteriaBacteriaLeukocytesLeukocytesLeukocyte, granular, andLeukocyte, granular, and

waxy castswaxy castsRenal tubular epithelialRenal tubular epithelial

cell castscell casts

pHpHpHpH

TraceTrace

PositivePositive

Common Findings inCommon Findings in::Nephrotic SyndromeNephrotic SyndromeMicroscopic:Microscopic:Oval fat bodiesOval fat bodiesFatty castsFatty castsWaxy castsWaxy casts

Microscopic:Microscopic:Oval fat bodiesOval fat bodiesFatty castsFatty castsWaxy castsWaxy casts

pHpHpHpH

++++++++

Eosinophilic CystitisEosinophilic Cystitis

Microscopic:Microscopic: Numerous eosinophilsNumerous eosinophils

(Hansel’s stain)(Hansel’s stain) NO significant castsNO significant casts..

Microscopic:Microscopic: Numerous eosinophilsNumerous eosinophils

(Hansel’s stain)(Hansel’s stain) NO significant castsNO significant casts..

pHpHpHpH

Urothelial CarcinomaUrothelial CarcinomaMicroscopic:Microscopic: Malignant cells on Malignant cells on urine cytology urine cytology (urine sample should(urine sample should be submitted be submitted separately toseparately to cytology, cytology, void or 24 hrs.)void or 24 hrs.)

Microscopic:Microscopic: Malignant cells on Malignant cells on urine cytology urine cytology (urine sample should(urine sample should be submitted be submitted separately toseparately to cytology, cytology, void or 24 hrs.)void or 24 hrs.)

pHpHpHpH

Bacterial Cystitis:

Urinalysis often shows increased protein and hemoglobin

Increased numbers of WBC, RBC, and/or bacteria are consistent with cystitis.

Urine Culture

Purpose:To identify the specific infectious agent.Antibiotic sensitivity test.

Urine Culture

Media Descriptions:C.L.E.D. (Cystine Lactose Electrolyte

Deficient Agar) is a non-selective medium that supports the growth of Gram (+) and Gram (-) species, specifically for enumeration of bacteria in urine.

URINALYSIS REPORT:

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