Blood Glucose TEST

A fasting test is performed after patient abstained from food/liquid for 12 hours prior to test

fasting blood sugar level of 99 mg/dL or lower is normal, 100 to 125 mg/dL indicates you have
prediabetes, and 126 mg/dL or higher indicates you have diabetes.

A random blood sugar is performed if patient isn’t fasting

Normal: less than 140 mg/dl. Prediabetes: 140–199 mg/dl. Diabetes: 200 mg/dl or above.

two hour post prandial blood sugar test

-screen for diabetes or effects of insulin dosage

normal range: less than 140 mg/dL 2 hours after eating

glucose tolerance test

-assessing insulin response to a glucose load

-high carbohydrate diet for 3 days, NPO after midnight before testing

Insulin-hormone produced pancreas in the islets of langerhans in the beta cells

diabetes risk factors- obesity , family history, high stress

diabetes type one: genetic condition that often shows up early in life

symptoms: nausea, abnormal thirst, constant urination rapid loss of weight

diabetes type two: mainly lifestyle-related and develops over time

symptoms: drowsiness, itching, blurred vision, tingling

*the three P's : polyuria, polyphagia, polydipsia

Glycogen: stored sugar

the action of sulfonylureas=stimulates insulin production in pancreas

HbA1c test-measures your average blood sugar levels over the past 3 months
-commonly used tests to diagnose prediabetes and diabetes

A normal A1C level is below 5.7%, a level of 5.7% to 6.4% indicates prediabetes, and a level of 6.5% or
more indicates diabetes.

how does the body glucose and it's main purpose?

metabolizes carbs to provide energy

LIVER FUNCTION TEST

What are the important functions of the Liver?

-Excretion, Plasma protein synthesis, Blood clotting, metabolism, detoxification

How can you assess the function of the Liver?

By performing a combination of biochemical tests known as Liver Function Tests (LFT)

-gives us an idea about the anatomical and physiological damage of Liver

What are some physical signs that can indicate a problem with the Liver?

Jaundice, Abdominal Pain & Fever, Stigmata of Chronic Cirrhosis, Complications of Liver Disease

What are the Liver Function Tests?

1) Test of Excretory Function

2) Test for Serum Enzymes
3) Test for Synthetic Function
4) Test for Metabolic Function

Tests for Excretory Function

Serum Bilirubin:
-TOTAL
-Conjugated
-Unconjugates

Serum Bilirubin and Hepatocellular Disorder

-Liver conjugates Bilirubin and excretes it into Intestines (bile)

-In patients with Hepatitis (Hepatocellular Disorder), Serum Total, Conjugated, and Unconjugated
Bilirubin levels are elevated

Serum Bilirubin and Cholestatic Disorder

-In Cholestatic Disease, serum Conjugated Bilirubin is predominantly elevated

-Biliary obstruction prevents conjugated Bilirubin from being excreted so its regurgitated into blood

Can you distinguish Hepatocellular disorder from Cholestatic disorder solely on Serum Bilirubin?

No. Serum Bilirubin mainly indicates the extent of damage.

in Cholestatic disease, Serum Conjugated Bilirubin is predominantly elevated

Can serum Bilirubin help indicate the extent of damage in Cholestatic Disorder?

Yes. The more serum conjugated Bilirubin, the more damage

Intrahepatic vs Extrahepatic Cholestasis

Intrahepatic Cholestasis:
-Inflammation of hepatocytes compresses on bile duct
Extrahepatic Cholestasis
-gall stone in common bile duct

What can be used as an indicator if the disease is predominantly resulting in Hepatocelluar damage OR
Cholestasis?

Enzyme Levels in Serum are a good indicator of whether the disease is predominantly resulting in
Hepatocellular damage or Cholestasis.

The degree of rise of serum enzymes corresponds to _____.

Extent of damage

What are some enzymes indicating Hepatocellular Injury?

Transaminases:
-ALT (Aminotransaminase)
-AST (Aspartatetransaminase)
Aminotransferases are raised in Viral Hepatitis, Drug Induced Hepatitis, and long standing obstructive
Hepatitis

ALT & AST levels in:

Acute Hepatitis: ALT>>>>AST
Alcohol Cirrhosis: AST>>>ALT

*2 to 1 ratio*

Enzymes Indicating Cholestasis (Both Intra and Extrahepatic)

ALP (also raised in growing children and pregnant women)

GGT
5-Nucleotidase

Where else can you find GGT? Gamma-glutamyl Transferase

Bone, Placenta (non hepatic)

How can you distinguish if Cholestasis is an indictor for Hepatic or Non Hepatic?

GGT is elevated if it's non hepatic cholestasis

What if there is an elevation of ALP with no elevation of GGT?

Elevation of ALP is non-hepatic

bone disease, placental ALP

What does it indicate if there is an elevation of GGT and no increase of ALP?

-Recent Alcohol use or ingestion of drugs that induce GGT synthesis

What is another indicator besides ALP and GGT for Biliary Disorders?

5'-Nucleotidase

Why is an ultrasound performed when ALP and GGT levels are indicated?
(Intra vs Extra Hepatic)

An abdominal ultrasound is performed to assess the Biliary Tree:

-BIliary Tree is Dilated: Extrahepatic

-Biliary Tree NOT Dilated: Intrahepatic

How can you test for the Synthetic Function of Liver?

-Serum Albumin + Globulins

Serum Albumin and Hepatitis

Serum Albumin levels are usually normal in ACUTE Hepatitis

Decreased in Serum Albumin level is an indicator of LONG staring Liver Disease

What is the only Globulin protein not synthesized by liver?

Gamma Globulins

What do elevated gamma globulin levels indicate?

Cirrhosis

What clotting factors are made by Liver?

F5, F7, F9, F10

What does increased prothrombin time indicate?

Increased Prothrombin Time indicates deficiency of clotting factors synthesizes by liver

Prothrombin Time and Cholestasis & Hepatocellular Disorders

Prothrombin Time is elevated in BOTH Cholestasis and Hepatocellular Disorders:

Cholestasis: Impaired absorption of Fat Soluble Vitamins including Vitamin K because of reduced entry
of Bile Acids to Intestines

*** Decreased Vitmamin K= reduced synthesis of Clotting Proteins

INR is used for the assessment of _____ disorders

-Clotting

When is it important to monitor Prothrombin Time?

If a Liver Biopsy or Surgery is planned

What Tests are used to evaluate metabolic function?

Blood Glucose
-Glycogen Synthase, Glycogenolysis
-Gluconeogenesis

Blood Ammonia
-Urea Cycle

Lipid Metabolism "fatty liver"

How does damage of hepatocytes by ethanol result in a fatty Liver?

Damage of Hepatocytes by ethanol results in fatty liver. Contributing Factors:

-Decreased B-oxidation due to increased NADH/NAD+
-Increased rates of FA synthesis due to increased Acetyl CoA levels
-Decreased rates of VLDL secretion due ot hepatotoxic action of ethanol

What are special test used to identify the cause of the altered LFT?

Serum AFP
Serum Iron, Transferrin and Ferritin
Serum Ceruplasmin
Serum Alpa1-AT

AFP Test -AFP is used as a TUMOR marker in patients with Liver Cancer

Serum Iron, Transferrin, and Ferritin

-Used in patients suspected of having Hemochromatosis

Serum Ceruplasmin

-Used in patients suspected to have Wilson's Disease (copper accumulation)

Serum Antitrypsin

Used in patients suspected to have inherited Alpha1-AT deficiency

When do we use Liver Function Tests (LFT's)?

-Assess the intensity of type of Jaundice

-Assess extent of damage to Hepatocytes
-Differentiate between Chronic and Actute
-Assess metabolic functions of Liver
-Monitoring Drug therapy of Hepatotoxic drugs
-Follow up on Liver disease
What are serum markers of Acute Hepatocellular Damage?

ACUTE: Hepatitis due to Viral or Drug Induced..

-elevated serum total Bilirubin
-elevated Conjugated Bilirubin
-Urine: Organe (positive for Con Bil)
-ALT + AST Elevated
-ALP and GGT elevated (Intrahepatic Cholestasis)
-Serum Albumin: LOW (Actute)
-Prothrombine Time elevated (decreased capacity to synthesize clotting factors)

What are serum markers of Cholestatic Jaundice?

-Serum Total Bilirubin: elevated

-Serum Unconjugated Bilirubin: elevated
-Urine Bilirubin: positive
-Urine Urobilinogen: ABSENT**
-ALT and AST: Elevated
-ALT and GGT: Elevated
-Serum Albumin: Normal
-Prothrombin Time: Elevated (decreased Vitamin K absorption; reduced clotting factor synthesis)

What are serum markers of Chronic Alcohol Cirrhosis?

Serum Bilirubin: Elevated

-Serum Conjugated Bilirubin: Elevated
-Albumin: Elevated ***
-Serum Globulin: Elevated
-AST>>>ALT (2 fold) Alcohol Cirrohsis**
-ALP: Elevated
-GGT: Elevated (induction by Alcohol)
-Prothrombin Time: Elevated
RENAL FUNCTION TESTS

Purposes of renal function tests:

-To evaluate glomerular filtration, tubular reabsorption, tubular secretion, and renal blood flow

Purpose of glomerular filtration clearance tests

-To measure the rate at which the kidneys can remove a filterable substance from the blood

Substances used to measure glomerular filtration rate:

Creatinine, beta-2 microglobulin, cystatin C, radioisotopes, exogenous procedure (requires an infused

substance), endogenous procedure (substance already present in the body)

Creatinine Clearance Test

Test to determine the amount of creatinine in mL completely cleared from the plasma in 1 minute.
Report in mL/min. Requires the urine volume in milliliters per minute (V), urine creatinine in mg/dL (U),
plasma creatinine in mg/dL (P).

Normal values of creatinine clearance:

Men: 107 - 139 mL/min

Women: 87 - 107 mL/min
Lower values in older people

Normal reference range of plasma creatinine: 05 to 1.5 mg/dL

How is creatinine produced in the body? Muscle destruction

Estimated GFR:

Tests do not require collection of timed 24-hr urine specimens

Estimated GFR tests : Serum creatinine, cystatin C, and Beta-2 Microglobulin

Serum creatinine test

Uses Modification of Diet in Renal Disease formula (MDRD) to calculate estimated GFR. Does not require
body size/weight, uses variables such as serum BUN, serum albumin, and ethnicity. All calculations are
available from the laboratory computer

Cystatin C

Small protein produced by all nucleated cells. Gets filtered by glomerulus, absorbed by the renal tubules
back into the serum, and broken down. No cystatin C is secreted in urine. Serum levels of Cystatin C
directly reflect GFR. Usually measured using immunoassay procedures.

Beta-2 microglobulin
Small protein that dissociates from leukocyte antigens at a constant rate. Rapidly removed from blood
plasma by kidneys, secreted in urine. Measured by enzyme immunoassay. Useful in detecting decrease
in GFR, but unreliable in patients with immunological disorders.

Exogenous Method

Uses radioisotopes to measure the removal of injected isotopes from the blood plasma by the kidneys.

Tubular reabsorption tests

AKA concentration tests. Good indicator of early renal disease, measures renal ability to concentrate
salts and water. Baseline for determining concentration is 1.010 specific gravity of the original
ultrafiltrate. Accurate results require the control of patient fluid intake.

Osmometry/Osmolarity/Osmolality

Used to test renal concentrating ability. Measurement of total solute concentration in urine. Used to
analyze changes in colligative properties based on number of solutes. Freezing point, boiling point,
osmotic pressure, and vapor pressure can be analyzed with instrument.

Colligative properties

Freezing and boiling points, osmotic pressure, and vapor pressure

Clinical significance of measuring osmolarity

Evaluates renal concentrating ability to monitor course of renal disease, fluid and electrolyte therapy,
secretion and response to ADH, and to diagnose hyponatremia and hypernatremia.

Renal Concentration Test procedure for Osmolality

Overnight deprivation of fluid for 12 hours followed urine collection

Urine osmolality of ≥800 mOsm (3:1 urine to serum ratio) is normal, test can be discontinued

If urine osmolality is abnormal, fluid restricted for additional 2 hours and another urine and serum
collection for osmolality testing is performed.

Further studies are needed if tests continues to be abnormal using ADH injections to test for diabetes
insipidus

Normal ratio of urine to serum (U:S ratio) for controlled fluid intake

=3:1. Normal tubular reabsorption is indicated by urine osmolality of 3:1.

Why are ADH injections needed for continued abnormal osmolality tests?

Determines whether the failure to concentrate urine is caused by diabetes insipidus that occurs with
ADH production problem or response of kidney to ADH.

How to test for Diabetes Insipidus

After abnormal results are determined from initial fluid deprivation, inject patient with ADH,
collect urine and serum specimens at 2 and 4 hours, test for osmolality.
-Result of 1:1 ration or <800 mOsm indicate lack of response to ADH by convoluted tubules.
-Result of 3:1 indicates inability to produce adequate amounts of ADH

Normal serum osmolality reference range

275 to 300 mOsm

Normal urine to serum ratio (uncontrolled intake)

1:1

Normal urine osmolality reference range

Depends on fluid intake or exercise, so it is difficult to establish

Why is osmolarity more useful than specific gravity to test urine concentration?

Osmolarity simply takes into account number of particles dissolved in the urine. Specific gravity can be
influenced by larger molecules while being less concentrated.
LIPID PROFILE AND CARDIAC MARKERS

Risk factors for cardiac disease:

Smoking
high blood pressure
diabetes
obesity/overweight
a family history of early heart disease
high blood cholesterol (high total and high LDL)
low HDL cholesterol.

What is hypercholesterolaemia?

An excess of cholesterol in the blood stream

What is the most important blood test for risk assessment?

Lipid profile

What does The lipid profile measures ?

Cholesterol
HDL-cholesterol
LDL-cholesterol
Triglycerides

What type of blood samples can be used for screening measurement of total cholesterol

Non-fasting capillary or venous samples

What Other tests that help measure the risk of cardiovascular disease (CVD).

High sensitivity C-reactive protein (hsCRP)

Lipoprotein A (Lp(a))

The hs-CRP test measures CRP within the range which is seen in otherwise healthy people so what does
it distinguish between

Low normal levels from people with high normal levels

High normal levels of hs-CRP in otherwise healthy individuals can be used to predict what ?

Future risk of:

heart attack
stroke
sudden cardiac death
peripheral arterial disease (disease of arteries not in the heart)

What is a Lipoprotein A (Lp(a))

-Is a lipoprotein consisting of an LDL molecule with another protein (Apolipoprotein (a)) attached to it.

Lp(a) is similar to LDL-C but how does it vary ?

Does not respond to lowering LDL-C by diet, exercise, or most lipid lowering drugs.

What does a high level of Lp(a) mean ?

Since the level of Lp(a) is not easily altered it can be used to identify people who might benefit from
more 'aggressive' treatment of other risk factors.

What are the 5 types of Fredrickson Classified Lipid Disorders

1- increased chylomicrons
2- increased cholesterol without high trig's
3- variants in apoE abnormal IDL
4- increased VLDL
5- increased VLDL and chylomicrons

What causes low HDL

Insulin resistance and metabolic syndrome lead

Severe inflammation

High trigs

Liver disease can decrease apoA1 synthesis

How does a high amount of triglycerides lower HDL

-By transferring cholesterol to TRL's

How does Hypertriglyceridaemia effect risk of CAD

Increased risk mostly due to its inverse correlation with HDL

High chylomicrons linked to what ?

Pancreatitis

What are the Other cardiac risk markers and what are there links ?

Homocysteine- may be measured particularly if other risk factors low

MTHFR- genetic link to homocysteinaemia
Fibrinogen- acute phase protein
Apo A- linked to HDL levels
Apo B- linked to LDL levels
Apo E- linked to IDL levels also to risk of Alzheimers
What is the most commonly used Cardiac Biomarker ?

Creatine kinase MB isoenzyme or as a ratio CKMB/Total CK

What Muscle Proteins can be used as Cardiac Biomarkers ?

Myoglobin
Troponin biochemical gold standard for detecting myocardial necrosis

What are the Benefits of Cardiac Troponin Measurement

Specific to cardiac damage- only cause of blood increase

Highly sensitive- very low in normal subjects
Detects Myocardial infarction in patients which can not be diagnosed using CK or CKMB

Definition of Myocardial infarction

Evidence of ischaemia with at least one of the following: symptoms of ischaemia, ECG changes of new
ischaemia, development of pathological Q waves in the ECG, or an imaging evidence of new loss of
viable myocardium or new regional wall motion abnormality
ARTERIAL BLOOD GAS

What is an ABG?

-a measurement of the partial pressure of O2, CO2, pH, and HCO3 in arterial blood

Who normally draws this bloodwork? Where is it drawn from?

-Respiratory therapists draw for this profile most commonly from the radial, brachial, and femoral
arteries.

What is acidosis?

-It is characterized by an abundance of H+ ions in the arterial blood, with a pH of less than 7.35.

What are the acids?

H+, CO2, and H2CO3 (carbonic acid)

How are acids produced?

They are produced during metabolic processes.

How are acids excreted from the body?

Some exit via the lungs; others are buffered and excreted via kidneys.

What is alkalosis?

It is characterized by a decrease of H+ ions in the arterial blood, with a pH of more than 7.45.

What are the bases?

HCO3 (bicarbonate)

What is a buffer?

A chemical substance that reduces changes in systemic pH by either releasing or binding H+ ions. It is
considered to be a moment by moment regulation.

Why is CO2 considered a "potential acid"?

Because it forms carbonic acid when combined with H2O.

What are compensatory mechanisms?

Mechanisms that try to prevent large changes in pH and/or attempt to correct alterations in acid-base
balance; compensatory mechanisms ALWAYS alter the pH towards a normal level.

Give 2 examples of compensatory mechanisms.

Lungs and Kidneys

What is partial pressure?

Pressure exerted by a single gas.

What is the goal in the regulation of pH?

To maintain or restore body pH to a normal level - either through buffers or compensatory mechanisms.

What are some examples of buffers?

proteins, carbonic acid - bicarbonate, phosphate

How do the lungs function as a compensatory mechanism?

By expiration, in which CO2 is eliminated from the body, thereby reducing the amount of acids. (note
that the lungs cannot eliminate H+ ions)

How is additional expiration from the lungs initiated? What is the timeframe involved?

Through chemoreceptors in the brain. It occurs within 1-3 minutes of detected imbalance, and lasts 12-
24 hours.

How do the kidneys function as a compensatory mechanism?

In alkalosis : they excrete HCO3

In acidosis : they excrete H+ in addition to increasing production and retention of HCO3

What is the timeframe involved in kidney compensation?

It begins to work within approximately 24 hours and continues until pH is WNL

What is the normal range for pH?

7.35 - 7.45

What are the normal limits for PCO2?

35 - 45 mmHg

What are the normal limits for HCO3?

22 - 26 mEq/L

What is the normal range for PO2?

80-100 mmHg

What does a low PO2 indicate and what is the potential (respiratory) result?

A low PO2 indicated hypoxia and can result in hyperventilation.

ABG Interpretation : What does the relationship between pH and PCO2 indicate?

A respiratory problem

ABG Interpretation : What does the relationship between pH and PHCO3 indicate?

A metabolic problem
If pH and PCO2 are going in opposite directions, what does this indicate?

A respiratory problem

If pH and PHCO3 are going in the same direction, what does this indicate?.

A metabolic problem

What type of compensation is indicated if either pH and PCO2 or pH and PHCO3 are out of range (i.e.
only 2 components)?

Uncompensated compensation

What type of compensation is idicated if pH, PCO2, and PHCO3 are all out of range?

Partially compensated compensation

What type of compensation is indicated if the pH is WNL and PCO2 and PHCO3 are out of range?

Fully compensated compensation (note : need to determine whether pH is closer to acidosis or alkalosis
to determine respiratory vs. metabolic)

How does the brain respond to retained CO2 in the body?

The blood vessels dilate, resulting in confusion.

What are the characteristic values of respiratory acidosis?

low pH; high PCO2 (lungs to not rid the body of CO2, the excess of which binds to H2O to form H2CO3)

What is the cause of respiratory acidosis? What can lead to this condition?

CAUSE = HYPOVENTILATION
factors leading to this include : trauma, disease, meds, CNS depression
[CNS depression (affects respiratory center in the brain); narcotics/sedatives/anesthesia (slow breathing
rate); trauma that results in impaired respiratory function; pneumonia, atelectasis, pulmonary
edema/pulmonary embolism]

What is the respiratory compensation for respiratory acidosis?

an increase in respiration rate (unless CNS depression)

What are the renal compensations for respiratory acidosis?

excretion of H+ ions, and production and retention of HCO3

What are the characteristic values for respiratory alkalosis?

high pH; low PCO2

What is the cause of respiratory alkalosis and what is this secondary to?

CAUSE = HYPERVENTILATION secondary to

pain
anxiety/fear
fever/sepsis
CNS lesions

What is the renal compensation for respiratory alkalosis?

a decrease in production of HCO3 (at around 24 hours)

Is metabolic acidosis or metabolic alkalosis seen more often in a clinical setting?

metabolic acidosis

What are the characteristic values of metabolic acidosis?

low pH; low PHCO3

(note : this is usually secondary to significant health problems such as DKA and CKD)

What are some causes of metabolic acidosis?

*renal failure (decreased production of HCO3 as well as decreased excretion of H+ ions)

*severe diarrhea (increased loss of HCO3)
*lactic acidosis (increased production of acids)
*asprin toxicity (AKA : ASA Toxicity)
*starvation
*anaerobic metabolism secondary to hypoxia (remember cell injury leads to lactic acid production and
accumulation)

What are the respiratory compensations for metabolic acidosis?

and increase in respiration rate & depth (Kussmauls respirations = deep, rapid resp. pattern to blow off
CO2 - this is very characteristic)

What are the characteristic values for metabolic alkalosis?

high pH; high PHCO3

What are some causes of metabolic alkalosis?

*increased renal production of HCO3

*excess ingestion of alkali (antacids/baking soda)
*GI suctioning/vomiting/bulimia/diuretic therapy (all involve a loss of acids

What is the respiratory compensation for metabolic alkalosis?

decreased respiration rate

ROUTINE URINALYSIS

Test urine for Renal or urinary tract disease such as nephritis/ nephrosis, UTI's

Urine samples:

easy to obtain
good way to screen asymptomatic population for undetected disorders
can be used to monitor progress of disease and effectiveness of therapy

Urine composition:

affected by diet, activity, metabolism, endocrine function, and body position

Normal constiuents 95%- water and 5% solutes
Urea, organic and inorganic chemicals

Urine formed elements

-usually not part of the original ultra filtrate

their presence may indicate a disease process

Completely abnormal constituent

result of some physical or metabolic problem

bacteria, cellular cast, oval fat bodies, amino acids, products of abnormal metabolism

May indicate a disease process

RBC

WBC

epithelial cells (renal/ transitional/ bladder/ squamous)

Hyaline cast/ granular cast/ Cellular cast/ crystals/ mucous/ bacteria, parasites, and yeast

Urine sediment

analyzed in a hemacytometer an individual elements are reported in 24 hours

Bence Jones protein

a urine protein with patients suffering from multiple myeloma

Urine collection
Chain of custody

Proper collection, labeling, handling must be documented from the time specimen is collected and
received
standardized form always accompanies specimen
collectors should be properly trained
specimen must withstand scrutiny- pre-employment
sport figures
military
probation

Urine Collection methods 2 types

Mid-stream- the patient begins voiding in the toiled then inserts specimen container in to the continuing
urine stream until the cup is halfway filled

Clean Catch- prior to voiding process the patient performs a series of steps to clean the external genital
tissues in effort to remove contaminating bacteria

Urine Collection- pediatric methods

baggie method is Okay for specimen collection

suprapubic aspiration /cystocentesis

-urine is obtained from a needle through the abdominal wall

-bacterial cultures (anaerobic cultures) for cytology

catheterized specimen

collected from a hollow tube threaded up the urethra into the bladder

for cultures or patient can't void

Ureteral catheterization- specialized catheterization to obtain samples from each ureters

Collection time of Urine Random

Random- collected at anytime most common but not accurate, can be affected by diet and physical
activity

First voided/ first morning specimen- recommended specimen for routine UA most concentrated
(pregnancy test preferred) and most likely to reveal abnormalities

Fasting must be done for diabetic monitoring

2 or 3 glass urine (prostatitis specimens) voiding process is divided into two- three segments

Collection time or Urine- timed

2, 12, 24 hr specimens patient is given very detailed instructions required for quantitative chemistry
tests

Urine specimen container

chemically clean- no contamination preferably sterile, disposable

pediatric: plastic bags with adhesive

tight fitting lid

clear plastic- ideally for routine urinalysis

non-routine and 24 hour collection, brown or dark colored containers to keep out light

Properly label the specimen- name, date, time of collection, hospital#, doctor delivered to lab
immediately

Preservation of the Urine specimen

Specimen Integrity

Test within 2 hours of collection or refrigerate if not

specimens can deteriorate

ketones can evaporate

bilirubin& urobilinogen destroyed by light if they are not in a dark container

Bacteria can multiply- metabolize/ use uf available glucose

modify urea molecule- resulting in release of ammonia- which makes pH increasingly alkaline

alkaline environment destructive

Specimen rejection

problems with patient/ specimen ID not labeled requisition and specimen labels do not match sample
collected on wrong patient

sample contaminated

QNS- improperly collected/ preserved

delay in transport

Labs have specimen rejection policies follow protocol

3 Classifications of Urine tests

Screening- detects presence or absence of a substance

positive or negative

Qualtitative (semiquantitative)- provides a rough estimate of the amount of the substance: usually
reported as neg, trace, 1+,etc....

Quantitative- accurate determination of the substance being detected: reported as specific amt
per/specific time or volume. ie mg/dL or g/24 hours

Addis count

accurate count to assess urine sediment

Best option to test urine

test urine within 1-2 hour

refrigerate 4 degrees C- ASAP following collection

Refrigeration will increase specific gravity and cause the precipitation of amorphous crystals

Dipstick testing of cold specimens reduce speed of reactions - can lead to erroneous results

Urine must be tested at room temperature

Freezing destroys formed elements but preserves bilirubin, urobilinogen, and porphobilonogen.
rarely used chem preservatives for routine urinalysis

Toulene- preserves chemical constituents, prevents bacterial mulitplication

Formalin- kills bacteria preserves the sediment but affects chemical tests

Thymol crystals- interferes with acid precipitates test for protein

Boric acid may cause crystal precipitation doesn't inhibit bacteria well

chloroform- inhibits bacterial growth but changes the characteristics of the cellular sediment

C&S transport kit - increase specific gravity and protein decreases pH.

Routine urine - organic

Uric acid- from purine catabolism

Urea- from protein and amino acid metabolism

Creatinine- by product of muscke metabolism

Testing for Urea and Creatine can be used to identify a fluid as being urine

Routine urine- Inorganic

anions- negative charge Cl, phosphate, sulfate

Cations- positive charge Na, K ammonium

small or trace amounts

ROUTINE FECALYSIS

Specimen collection of Stool

• Clean, dry, wide-mouthed containers

• Sealed and sent to the laboratory immediately after collection.

• Preserved specimens can usually be kept at room temperature

- stool should not be contaminated with urine

Large gallon containers

for 24-, 48-, and 72-hour stool collections for fecal fat and urobilinogen; specimens are normally
refrigerated throughout the collection period

- also used for parasitic diagnostic tests, if antiparasitic drugs are working properly

Reasons for rejecting stool specimen

• Specimens in unlabeled containers

• Nonmatching labels and requisition forms

• Specimens contaminated with urine or toilet paper

• Containers with contaminated exteriors

• Specimens of insufficient quantity

• Specimens that have been improperly transported

Routine stool examination

- Physical Analysis

- Chemical Analysis

- Microscopic Analysis

physical examination

Consistency

Color

Gross abnormalities
Consistency

- Degree of moisture in a stool specimen

- Formed, semiformed, soft, loose, or watery

stool

- formed in large intestine

- water can either be reabsorb or released

- to check consistency, open the lid slightly enough for applicator stick to enter

stercobilinogen

causes normal brown color of stool

Color

- The normal color of the stool is brown.

Black color stool

- upper GIT bleeding

- Iron therapy

- Charcoal used in black foods (non-patho)

- Bismuth (antacids)

Red color stool

- Lower GIT bleeding

- ingestion of beets and food coloring

- Rifampim

Green color stool

- Biliverdin (abnormal presence of liverdin)

- oral antibiotics

- green vegetables

Pale yellow, gray, white color stool

- bile duct obstruction

- barium sulfate

presence of mucus, blood steak - brown color

- dysentery

- malignancy

Gross abnormalities

- macroscopic structures, organisms seen in the stool by the naked eyes

- Adult worms, proglottids and parasitic indications (pus and mucus)

- Samples containing adult worms may be carefully washed through a wire screen. This process allows
for the retrieval and examination of the parasites for identification purposes.

proglottids

eggling segments of tapeworm

Chemical Examination

detection of occult blood (hidden blood)

- screening for possible colorectal cancer

- prompts physicians to do further testing

- other chem examinations: fat content, urobilinogen, proteins, etc

Hema-screen Guiac Slide Test Kit

1. Hema-Screen Slide

2. Hema-Screen developer (<6% H2O2 )

Hema-Screen developer

contains hydrogen peroxide

FOBT Result
(+) any trace of blue coloration
(-) no detectable blue coloration
False-Positive FOBT
• Aspirin and anti-inflammatory medications
• Red meat (beef and goat)
• Horseradish
• Raw broccoli, cauliflower, radishes, turnips
• Melons
• Menstrual and hemorrhoid contamination
False-Negative FOBT
• Vitamin C >250 mg/d
• Iron supplements containing vitamin C
• Failure to wait specified time after sample is applied to add the developer reagent
Microscopic Examination
- To detect the presence of parasites in a stool specimen
- It can also reveal an artifact (non-patho) caused by food
- It should be performed on a fresh specimen
- for parasitology

Microscopic Examination Procedures

1. Direct wet preparations
2. Concentrated technique
3. Permanently stained smear

NOPS
No ova and/or parasite seen
NIPS
No intestinal parasite seen

Artifacts, Pseudoparasites, and Confusers

- White blood cells
- Red blood cells
- Macrophages
- Charcot-Leyden crystals
- Epithelial cells
- Eggs of arthropods, plant nematodes, and other spurious parasites
- Fungal spores
- Elements of plant origin
- Plant and animal hairs
- Muscle fibers
- Fat globules

White blood cells

appears as circular in structure, with more than 1 nuclei inside and granules structures
inside it

Red blood cells

appears circular in structure with no nucleus and granules inside it
- appears cleaner than WBC

Epithelial cells
- very large irregularly shaped cells
- covers the entire view of the microscope
Charcot-Leyden crystals
often the products of metabolism of certain antibiotics
- appears spindle shape / needle like crystalline structure

Macrophages
appears larger than WBC and RBC

Eggs of arthropods, plant nematodes, and other spurious parasites

- most probably accidental ingested during meals (manner of food preparation)

fungal spores
- looks like RBC
- circular and clean inside
- looks like they are dividing under microscope
- budding appearance

fat globules
appears circular and are often yellow colored cells and shiny like oil in water

muscle fibers
high protein diet

Direct fecal smear

1. About 2 mg of stool (amount forming a low cone at the tip of an applicator stick)
2. 0.85% sodium chloride solution (NSS) - appears clear
3. Cover with cover slip

Cover with cover slip

- Primary useful in the detection of motile protozoan trophozoites (pale and transparent).
- to avoid contamination of microscope

Weak iodine solution (Lugol's and D' Antoni)

- temporary stain to demonstrate nuclei
- Helminth eggs and larvae can be detected using this preparation

Helminth eggs Cytoplasm

Golden yellow

Helminth eggs Nucleus

Pale and refractile

Helminth eggs Glycogen

Deep brown
FECAL OCCULT BLOOD TEST
purpose of FOBT
Qualitative method for detecting occult blood in the stool

May be indicative of asymptomatic GI diseases such as:

- Colorectal cancer
- Polyps
- Colitis

foods recommended for FOBT

Starting 2 days before testing and continuing throughout test period, patient should eat:
1. No red meat
2. High residue diet

foods to avoid for FOBT

During test period, patient should avoid raw fruits and vegetables containing
peroxidase-like substances, including:
1. Turnips
2. Broccoli
3. Horseradish
4. Cauliflower
5. Cantaloupe
6. Parsnips
7. Red radish

materials needed for FOBT

1. Henry Schein One Step slides
- Use special electrophoresis paper impregnated with guaiac resin
- Contains positive and negative performance standards

2. Henry Schein One Step developing solution

- Mix of hydrogen peroxide and denatured ethyl alcohol
- In aqueous solution

3. Laboratory pack
- Instructions for use
- 100 single slides with performance standards
- Two 10 mL bottles of developing solution
- 100 applicator sticks

4. Patient pack
- Instructions for use
- 150 patient slides with performance standards
- Three 10 mL bottles of developing solution
- 150 applicator sticks
- Patient instructions
- 50 foil-lined postal-approved mailing pouches

5. Clock/timer
- Time at least 2 minutes

storage conditions of FOBT

1. Store test slides and developer at room temp (59-86 F/15-30 C)

2. DO NOT refrigerate or freeze

3. Protect from heat, humidity, and light

4. DO NOT store with volatile chemicals

5. DO NOT use discolored slides (normally beige in color)

6. Keep developing solution tightly capped when not in use

benefits of storing slides as recommended

Maintain sensitivity up to 3 years from manufacture date

what can occur if slides are not stored as recommended

1. Discolor and turn blue

2. Lose sensitivity

benefits of storing developing solution as recommended

Remains stable for at least 3 years from manufacture date

internal quality control for FOBT

Positive and Negative Performance Monitors

Must be performed and documented for all individual patient tests performed

Results can only be reported when both Positive and Negative Performance Standards
give expected results

Corrective action must be performed and documented whenever Positive and Negative
Performance Monitors fail to give expected results

results of FOBT
Any trace of blue = + test
No indications of blue = - test

Important that results be read 30-60 seconds after developing solution is applied
- Color reaction fades after 2-4 minutes
interpreting results from a blue discoloration on the guaiac test paper
Blue color migrates outward and forms a blue ring at the edge of the wetted area
- Considered negative result
- Guaiac paper around fecal smear will remain off-white in color

Any blue directly on the edge of the fecal smear

- Considered a positive result

Distinct green color with no blue

- Due to high bile content appearing at edge of smear
- Considered negative result

Blue/blue-green color
- Considered positive result

limitations of FOBT procedure

1. Results are designed for preliminary screening only
- Not intended to replace diagnostic tests (barium enema, proctosigmoidoscopy, X-rays,
etc.)
- Not considered conclusive evidence for presence/absence of GI bleed/pathology, as
GI cancers/adenomas do not always bleed

2. Some oral medications can interfere with results and should therefore be avoided 2
days prior to and during testing period
- Can cause GI irritation and occult bleeding in some patients (aspirin, corticosteroids,
reserpine, phenylbutazone, indomethacin
- Can cause false negatives (ascorbic acid)
- Can cause false positives (iron)

3. Patients with bleeding from other conditions should not be tested while conditions are
present
- Hemorrhoids
- Dental work
- Constipation
- Menstrual bleeding

4. DO NOT collect specimen if pt is using rectal preparations

MALARIAL MICROSCOPY
Microscopy remains the GOLD STANDARD for diagnosis
Typically thick and thin blood smears are stained with Giemsa stain

Thick smears > presence of parasites

Thin smears > species-level identification

Microscopy - blood smears

Ring-form trophozoites (rings) of P. falciparum are thin and delicate structures
visualized within the red blood cell

Rings may possess one or two chromatin dots

Multiply-infected RBCs are not uncommon

P. falciparum blood smear showing numerous ring-form parasites

blood smears (pic)

diagnosis
In ENDEMIC areas, patients may have POSITIVE smears and NOT have clinical
malaria

P. falciparum infected RBCs may be sequestered (cytoadherence) and NOT observed

in the peripheral blood

Estimate number of infected RBCs per μl of blood:

Poor prognosis if >500,000 parasites/ μl blood (~10% parasitemia).

Malaria - treatment
Chloroquine (for susceptible strains)
Quinine
Mefloquine (neurologic side effects)
Doxycycline (inexpensive, but tricky to use)
Atovaquone+Proguanil (easy to use, but expensive)
Artemisinin (new)
Primaquine

control of malaria
Insecticide treated bed nets (ITNs)

Indoor residual spraying - walls within homes

Vector Control - control/ eradication of mosquito population

SEMEN ANALYSIS

What are the tests performed on seminal fluid?

Liquefaction, volume, viscosity, pH, count, motility, morphorlogy and viability.

What is most of semen composed of?

viscous liquid that furnishes nutrients to the sperm.

What does the milky fluid from the prostate contain?

Acid phosphatase and enzymes which result in coagulation and liquefaction.

Why are plastic containers not recommended for sperm collection?

Plastic decrease the motility of the sperm.

How should the specimen be handled?

Keep the specimen at room temperature and get to the lab w/i one hour after collection.

What should happen to fresh semen after 30 min?

Clot and then liquefy, therefore in order to measure liquefaction accurately, a fresh specimen is needed.
None of the other tests can be performed on the sample until it liquefies.

What is normal volume?

2-5 mL.

How is viscosity rated?

watery = 0 to gel like = 4.

What is normal pH for semen?

7.2-8.0. A pH paper or dip reagent strip is used to get the pH on the sample.
What is a normal sperm count?

20-60 million sperm/mL of seminal fluid.

When analyzing, what should be measured first before diluting?

Analyzing the sample for motility and morphology due to diluting changes their morphology (water
causes their tails to kink up into tight curls).

How is the specimen diluted for a count?

1:20 ration and is counted on a hemocytometer.

Motility must be observed within how long after collection allowing for liquefaction?

w/i an hour.

What is normal motility?

50-60%

How do the sperm need to be moving?

In a straight line not a circle.

How many sperm will have abnormal forms incapable of fertilization?

30%

Where do the sperm come to full maturity at?

The epididymis

What's one of the stains that can be used on semen?

Wright stain (which is used to differentiate white blood cells).

How is morphology turned out?

Percentage of normal forms, percentage of abnormal forms and then describe the types and percentage
o abnormal forms present.

What is viability?

term used to denote when the patient has a normal sperm count, but the sperm present aren't alive.

How long can the post vasectomy patient take to achieve sterilization?

2 months.

What is normal sperm count?

>40 million

Describe how seminal fluid comes together during ejaculation.

Seminal fluid is composed of 4 factions made in different places. Most of the semen is composed of a
viscous liquid that furnishes nutrients to the sperm. Then the milky fluid from the prostate contains acid
phosphatase and enzymes that act on the fluid from the seminal vesicles resulting in coagulation and
liquefaction.

Describe the steps taken by the lab on a post vasectomy specimen.

Two months post-vasectomy, a drop of seminal fluid is put on a slide, coverslipped and look for sperm.
Not one sperm, dead or alive should be overlooked. Then the specimen is spun down and examine a
drop of the sediment to make sure we don't miss a sperm.
STAPHYLOCOCCAL IDENTIFICATION

3 Major Stapholococci

S. aureus, S. epidermidis, S. saprophyticus

Staph traits

Gram positive, halotolerant, found everywhere, pus-forming, opportunistic pathogens, cause skin
infections and septicemia

Most toxic staph

S. aureus - causes TTS

SM110 agar

for staph, allows us to see colors

Agar tests used:

SM110, DNase, Blood, Coagulase, MSA

alpha-hemolysis

partial red blood cell lysis. Causes GREEN color

beta-hemolysis

Complete red blood cell lysis. Zone of inhibition.

gamma-hemolysis

NO LYSIS

DNase Test Agar contains

Methyl green indicator

DNase Agar tests

DNA hydrolysis denoted by zone of inhibition (coagulase pos. ONLY have DNase)

S. aureus results

Yellow and ALL positive (DNase, mannitol, blood, coagulase)

S. epidermidis

White, ALL negative

S. saprophyticus

White, positive Mannitol fermentation, the rest negative

S. aureus has 2 proteins

Coagulase and Protein A

Agglutination latex beads have two antibodies

Fibrinogen (binds to coagulase) and IgG (binds to Protein A)

Agglutination Test

ONLY S. aureus agglutinates!! If negative result, it is NOT S. aureus

ACID FAST BACILLI TESTING

acid fast bacilli

form long, branching cells

w/in Order Actinomycetales

Mycobacterium most significant genus

Mycobacterium tuberculosis

principle cause of tuberculosis (TB) in humans

what is M. tuberculosis referred to as sometimes?

tubercle bacillus

why is M. tuberculosis so resistant to ordinary stains?

cell wall has high lipid content

what is TB highly resistant to

acids & bases

used to culture it

how long can it take TB to form visible colony when cultured?

3-5 weeks

very slowing growing org

what would happen to specimens if treated with strong base?

normal flora bacteria mixed w/ TB would be killed

TB symptoms

fluid accumulation in lungs--mimics pneumonia

lesions on lung--can solidify to form nodules

bloody sputum--lesions next to blood vessels causing it to perforate

TB disease process

inflammation at infected area

T cells, B cells, macrophages & C' active at infection site

tubercles

nodules on lung

what occasionally happens if host fails to mount immune response?

uncontrolled multiplication of TB

uncontrolled multiplication of TB can lead to

Miliary TB

Miliary TB

M. tuberculosis spreads throughout body via blood

TB transmission

inhalation--almost always

ingestion--rarely

source of M. tuberculosis via ingestion

contaminated milk--TB bact removed by pasteurization

can lead to intestinal TB

TB diagnosis

positive:
-skin test

-quantiferron--blood test

-chest x-ray--look for tubercle

-sputum specimen--culture, AFB stain, PCR

TB diagnosis via culture

inoculate special media--Lowenstein Jensen agar, special liquid media

incubate up to 8 weeks

TB rapid nonculture test

on sputum specimen--AFB smear, PCR

benefits--rapid

drawback--smears not as sensitive as culture, PCR expensive

host immune response results in

patient becoming hypersensitive to tubercle bacillus

what is the basis to the TB skin test

hypersensitivity

TB skin test

tuberculin test--Mantoux, Tine

inject patient w/ purified protein derivative (PPD) of org, wait 2 days

TB skin test possible reactions

positive-redness at injection site--hypersensitive--have sensitized T cells

negative--patient has no history of exposure

if patient has positive skin test next steps are to

run chest x-ray--looking for tubercles

collect sputum specimen--see if infection is active

TB vaccine

BCG vaccine

not widely used in U.S.--controversy over safety & effectiveness

TB treatment

antibiotics--tough, resistant strains readily develop

drug synergy--INH, ethambutol, rifampin

can last as long as year

MDRTB--problem, HIV patients

prevent TB

isolate & treat cases of active infection

pasteurize milk

Mycobacterium avium (MAI, MAC)

AFB infection normally seen in AIDS patients

where is M. avium commonly found

soil

water

associated w/ some animals

how does M. avium cause disease in immunocompetent patients

form of pneumonia

rarely causes disease in these patients

how does M. avium infect AIDS patients

invade & infect variety of tissues

blood, lung, GI tract

can be fatal

M. avium treatment

similar to TB

MDRMAI--becoming problem

Mycobacterium leprae

cause of leprosy

Hansen's disease

where has M. leprae been successfully cultured

in lab--in susceptible animals

armadillos, chimps, mice

Lepromatous

form of leprosy

cutaneous

characterized by:

-disfiguring nodules

-deformed hands & feet

-tissue destruction

Tuberculoid

less severe form of leprosy

involves nervous system

loss of feeling & skin pigmentation

when is leprosy usually acquired

during childhood

following prolonged contact--can run in families

leprosy incubation period

2-12 years

why is incubation period so long

bacteria has thick cell wall--protects it from hosts immune system, retards uptake of nutrients--causes
extremely slow growth

leprosy transmission

prolonged close contact--not highly contagious

how to diagnose leprosy

positive AFB stain

PCR from skin scrapings--detects bacteria's DNA

leprosy treatment

Dapsone--sulfa drug

Rifampin

how to control leprosy

segregation & treat cases

new culture methods--possible vaccine

Actinomyces

causes actinomycosis

actinomycosis

chronic disease

characterized by destructive abscesses of jaw & other parts of body

actinomyces characteristics

anaerobic

acid fast

can often be isolated from (apparently) healthy human oral cavity

can cause disease when person under trauma

how to diagnose actinomycosis

culture org from lesions

actinomycosis treatment

penicillin

cephalosporins

erythromycin

Nocardia

causes nocardiosis

AFB

can cause wound infections

where is nocardia often found

soil
how is nocardia acquired

inhalation

how does nocardia infect

starts in lung

can spread to other tissues via blood

forms lesions at infected tissues

nocardia can emerge as secondary infection in which susceptible people

ppl w/ other diseases

AIDS patients

nocardia treatment

sulfa drugs

industrial use for AFB

not used much

hard to grow
ENTEROBACTERIACAE TESTING

True or False.

Members of Enterobacteriaceae account for more than 70% of all reported urinary tract infections.

True.

Where in the environment are members of Enterobacteriaceae normally found?

They are ubiquitous world wide in the soil, water, and vegetation.

What are the common structural features that all members of Enterobacteriaceae share?

They all are:

Gram Negative Rods, non-spore forming, and have a common core polysaccharide component in their
LPS layer.

LPS is a common feature of all Gram Negative organisms.

Enterobacteriaceae are classified based on what structures/antigens?

All Enterobacteriaceae are classified according to:

1. Somatic O polysaccharides.

2. Capsular K antigens.

3. Flagellar H proteins.

What are the KEE organisms and what is unique about their ability to metabolize lactose when
compared to other Enterobacteriaceae?

Klebsiella, E. Coli, and Enterobacter family of organsims can vigorously ferment lactose to produce acidic
waste products and all produce gas.

What are Primary Pili (also called Fimbriae) and what are they used for?

Primary Pili are small projections that radiate out from the bacteria. They are used for adherence to host
cells and tissues, but not for locomotion.
True or False.

Enterobacteriaceae can undergo DNA scavaging from the environment, but cannot form sex pili to
directly exchange DNA.

False.

They can express sex pili if they posses the appropriate plasmid for it.

Somatic O Antigen's can be found as a component of what structure in Enterobacteriaceae?

O ag's are the outter most component of the LPS layer.

What are the common biochemical characteristics of all Enterobacteriaceae?

They are all:

1. Facultative Anaerobes.

2. Glucose fermenters.

3. Catalase Positive.

4. Oxidase Negative.

5. Reduce Nitrate to Nitrite. (NO3- to NO2-)

True or False.

Citrobacter is not capable of fermenting lactose and therefore is not included in the KEE organisms.

False.

Citrobacter can ferment lactose, just at a very slow rate.

Which members of Enterobacteriaecae are not motile?

Klebsiella. Yersinia, and Shigella. (KYS, kiss)

In order the of U-IMViC, what are the test results for Citrobacter?

UIMViC = +, −, +, −, +

(Urease: Positive.

Indole: Negative.

Methyl Red: Positive.

VP: Negative.

Citrase: Positive.)

In order the of U-IMViC, what are the test results for Enerobacter supp?

UIMViC = −, −, −, +, +

(Urease: Negative.

Indole: Negative.

Methyl Red: Negative.

VP: Postitive.

Citrase: Positive.)

In order the of U-IMViC, what are the test results for E. Coli?

UIMViC = −, +, +, −, −

(Urease: Negative.

Indole: Positive.

Methyl Red: Positive.

VP: Negative.

Citrase: Negative.)

In order the of U-IMViC, what are the test results for Klebsiella pneumoniae?

UIMViC = +, -, -, +, +

(Urease: Positive.

Indole: Negative.

Methyl Red: Negative.

VP: Positive.

Citrase: Positive.)

In order the of U-IMViC, what are the test results for Proteus Mirabilis?

UIMViC = +, -, +, −, ±

(Urease: Positive.

Indole: Negative.

Methyl Red: Positive.

VP: Negative.

Citrase: Variable.)

In order the of U-IMViC, what are the test results for Serratia Marcescens?

UIMViC = -, -, -, +, +

(Urease: Negative.

Indole: Negative.

Methyl Red: Negative.

VP: Positive.

Citrase: Positive.)

In order the of U-IMViC, what are the test results for Pseudomonas Aeruginosa?

UIMViC = -, -, -, -, +

(Urease: Negative.

Indole: Negative.

Methyl Red: Negative.

VP: Negative.

Citrase: Positive.)

Based on growth on media and biochemical tests, what is the easiest way to identify an unknown as
Pseudomonas Aeruginosa?
P. Aeruginosa does not ferment either glucose or lactose and will change the pH of a media to basic in
nature.

U-IMVC is only positive for Citrase and it is Oxidase positive.

No Enterobacteriaceae are oxidase positive.

Based on essentially identical U-IMViC results for Citrobacter and Proteus Mirabilis by what other means
can you differentiate them?

Both Citrobacter and Proteus are UIMViC +, −, +, −, +.

The key distinguising feature is that Proteus is strongly Urease positive and Citrobacter is only weakly
urease positive.

Citrobacter is also a slow lactose fermenter and gas producer where Proteus is neither. (Both produce
H2S)

Based on essentially identical U-IMViC results for Enterobacter and Serratia, by what other means can
you differentiate them?

Both are UIMViC −,−,−,+,+.

Key difference:

Enterobacter is a KEE organism and vigorously ferments lactose.

An unknown bacteria has an U-IMViC of −,−,−,+,+. What are the possible organisms and what test or
media would you use to distinguish?

Both Enterobacter and Serratia share this UIMViC result sequence.

Use EMB or MAC agar to check for vigorous lactose fermentation. A positive fermentation result
indicates Enterobacter.

An unknown bacteria has an UIMViC of +, -, +, -, +. What are the two possibilities for organism and what
tests will allow you to positively identify which of the two it is?

Both Citrobacter supp. and Proteus Mirabilis share this UIMViC sequence.

Citrobacter will slowly ferment lactose and produce non-sulfurous gas. Proteus Mirabilis will not do
either.

To identify, grow bacteria on MAC or EMB, Use Kligler Iron Agar, or simply check to see if Urease activity
was exceptionally vigorous indicating Proteus.
COMPLETE BLOOD TEST

What are the components of the CBC?

WBC, RBC, platelets, Hemglobin, Hematocrit, RBC indices (MCV, MCH, MCHC), RDW, MPV

(may also include a WBC differential)

What is the normal range for the WBC count?

5,000 - 10,000 /ul

What are some possible reasons for leukocytosis?

Acute infection (bacterial)

Pregnancy

Leukemia

What are some possible reasons for leukopenia?

Chemotherapy

Viral infection

Aplastic anemia

Instead of solely looking at the WBC count, what must you consider when evaluating the hematologic
status of a patient?

You must consider the individual absolute counts of each leukocyte type rather than the total WBC
count. For such a critical evaluation, the first step is to order a CBC with differential.

What is the normal range for the RBC count?

~4-6 million

Female: 4.2 - 5.5 million/ul

Male: 4.6 - 6.2 million/ul

What are some possible reasons for high RBC count (polycythemia or erythrocytosis)?
Polycythemia vera

Dehydration

High altitude

What are some possible reasons for low RBC count (anemia)?

Blood loss (acute or chronic)

Leukemia

Nutritional Deficiency

Hemolysis

What is the normal range for hemoglobin (Hgb)?

Female: 12-16 g/dL

Male: 13-17 g/dL

(The normal range of Hgb is highly age & sex-dependent, with men having higher values than women &
adults having higher values than children - except neonates, which have the highest values of all.)

What is hemoglobin?

the oxygen transporting component of the blood.

Carries oxygen to the tissues & carbon dioxide back to the lungs.

What is the normal range for hematocrit (Hct)?

Female: 38-47%

Male: 40-54%

What is hematocrit?

also known as the packed cell volume or PCV. It is a measure of the total volume of the erythrocytes
relative to the total volume of whole blood in a sample.
What are some possible reasons for a high hemoglobin/hematocrit?

Same as high red cell count:

Polycythemia vera

Dehydration

High altitude

What are some possible reasons for a low hemoglobin/hematocrit?

Same as low red cell count:

Blood loss (acute or chronic)

Leukemia

Nutritional Deficiency

Hemolysis

What are the red blood cell indices?

Mean Cell Volume (MCV)

Mean Cell Hemoglobin (MCH)

Mean Cell Hemoglobin Concentration (MCHC)

(Hemoglobin, Hematocrit, & RBC count are used to mathematically derive the erythrocyte indices.)

What does the MCV measure & what is considered normal?

Measures average red cell size.

Normal: 80-100 femtoliters

MCV > 100 fl = macrocytic

MCV < 80 fl = microcytic

What does the MCH measure?

Measures average weight of hemoglobin in red cells

What does the MCHC measure & what is considered normal?

Measures average concentration of hemoglobin in red cells.

Normal: 32-36%

MCHC<32% = hypochromic

MCHC>36% = laboratory error or red cells are spherocytes

What is the normal range for the Platelet Count (Plt)?

Normal range: 150,000 - 400,000

(One of the first lines of defense in helping blood to clot)

What are some possible reasons for a high platelet count (thrombocytosis)?

Post-splenectomy

Polycythemia vera

Acute blood loss

What are some possible reasons for a low platelet count (thrombocytopenia)?

Aplastic anemia

ITP & TTP

DIC

What does the RDW measure & when is it useful?

Red Cell Distribution Width = a numerical expression which correlates with the degree of anisocytosis
(variation in size of the population of red cells).

The RDW may be useful in monitoring the therapy for iron deficiency anemia or megaloblastic anemias.
As the patient's new, normally sized red cells are produced, the RDW initially increases, but then
decreases as the normal cell population becomes the majority.

What does the MPV measure?

Mean Platelet Volume = a numerical expression of platelet size

What is the differential?

A process where 100 white blood cells are differentiated. The differential also includes RBC, WBC, &
platelet morphology.

What are the six normal white blood cells seen in peripheral blood?

Band neutrophil (1-5%)

Segmented neutrophil (50-70%)

Lymphocyte (20-40%)

Monocyte (1-10%)

Eosinophil (1-3%

Basophil (0-1%)

What is the primary function of neutrophils?

phagocytosis

What is the term for an increased number of neutrophils & what are possible causes?

Possible causes for Neutrophilia:

bacterial infection

leukemoid reaction

What is the term for a low number of neutrophils & what are possible causes?

Possible causes for Neutropenia:

viral illness

thyroid disorders

aplastic anemia

What does "a shift to the left mean"?

What does "a shift to the right" mean?

A large number of immature (band) neutrophils is called "a shift to the left."

A large number of segmented neutrophils is called "a shift to the right."

What are eosinophils?

Pro-inflammatory cells that are capable of either protecting or damaging the host depending on the
situation. Their granules are toxic to parasites.

What is the term for an increased number of eosinophils & what are possible causes?

Possible causes for Eosinophilia:

allergic disorders

parasitic infections

What is the term for a low number of eosinophils & what are possible causes?

Possible causes for Eosinopenia:

shock

trauma

surgery

What is the primary function of basophils?

to enhance the immuno-inflammatory reaction

What is the term for an increased number of basophils & what are possible causes?

Possible causes for Basophilia:

allergic reactions

What are the major functions of lymphocytes?

Major function depends on the type of lymphocyte. T-cells are involved in cellular immunity & B-cells
are involved in humoral immunity (antibody production).

What is the term for an increased number of lymphocytes & what are possible causes?

Possible causes for Lymphocytosis:

viral infections

TB

CLL (chronic lymphocytic leukemia)

What is the term for a low number of lymphocytes & what are possible causes?

Possible causes for Lymphocytopenia:

AIDS

Hodgkin's lymphoma

What are reactive or "atypical lymphs"?

Occasionally lymphocytes will look a little different when they are responding to various stimuli. These
are called reactive or "atypical lymphs" which are commonly seen during viral disorders.

What is the primary function of monocytes?

phagocytosis

What is the term for a high number of monocytes & what are some possible causes?

Possible causes of Monocytosis:

recovery phase of an acute infection

What does the reticulocyte count measure & what does it indicate?

measures the % of reticulocytes (slightly immature RBCs) in blood. The number of reticulocytes in the
blood indicates how quickly they are being produced & released by the bone marrow. Normally, the
body will respond to bleeding or hemolytic anemia by an increased rate of RBC production.

What is the Erythrocyte Sedimentation Rate (ESR) test?

A simple test used to determine the presence of inflammation. This is not a diagnostic test but can be
used to monitor disease therapy.

What is a bone marrow exam used to evalute?

Evaluates various hematopoietic disorders such as leukemia, lymphomas, & certain anemias. Also used
to determine if a leukemia is in remission following various therapy options.
ERYTHROCYTE SEDIMENTATION RATE

State the principle of ESR.

Measures settling of erythrocytes in diluted human plasma in a 1 hour period. Used clinically to indicate
inflammation, differentiate various diseases, or to monitor therapies.

What is the normal range for ESR in men?

0-15 mm/hr

What is the normal range for ESR in women?

0-20 mm/hr

State the specimen requirement for ESR and time limits for performing the test.

Specimen should be whole blood anticoagulated with EDTA. Fresh blood at room temperature should be
tested within 2 hours; if refrigerated, within 6 hours.

For how long do you time a sed rate?

1 hour

State at least four sources of error when performing and ESR.

1. Incorrect blood:diluent ratio.

2. Bubbles in the Westergren tube.

3. Tilting of the tube, which accelerates the fall of erythrocytes.

4. Vibration, for example from a nearby centrifuge.

5. Age of specimen (not used within 2 hours at room temp; within 6 refrigerated)

6. Temperature (must be between 20-25 degrees Celsius and blood must be room temp.)

The ESR can be used to differentiate appendicitis from ___________.

acute pelvic inflammatory disease or ruptured ectopic pregnancy (ESR normal in appendicitis)
The ESR can be used to differentiate angina pectoris from:

myocardial infarction (ESR normal in angina pectoris)

Name various factors which affect the ESR.

RBC size and shape

plasma fibrinogen

globulin levels

mechanical/technical factors

The ESR is directly proportional to _________.

RBC mass

The ESR is inversely proportional to ___________.

plasma viscosity

What method is used in ESR? Why?

Westergren method; greater distance of sedimentation measured in the longer Westergren tube.

How much mL of blood is required for ESR?

1 mL

What method was previously used for ESR?

Wintrobe

Can hemolyzed specimens be used in ESR?

No!

When reading the distance of the ESR, should the buffy coat be included?
No

Is the ESR increased or decreased in sickle cell anemia?

decreased

Is the ESR increased or decreased with an increased room temp?

increased

Is the ESR increased or decreased when the ESR rack is unleveled?

increased

Is the ESR increased or decreased when a nearby centrifuge is in use?

increased

Is the ESR increased or decreased with elevated plasma immunoglobulin?

Increased
PT AND APTT

What are the 3 main steps in the formation of a clot?

Constrict blood vessel in area of the hole, form a platelet plug, and seal the plug with fibrin.

The time required for the formation of a fibrin clot when plasma is added to a thromboplastin-calcium
mixture is called?

Prothrombin time (PT)

The PT test is a measure of the extrinsic and common pathways of coagulation involving factors?

III, VII, I, II, V, and X

(Rate of fibrin formation depends on the levels of these factors)

The PT test is a valuable screening procedure used to indicate possible factor deficiencies of what
pathway/s?

Extrinsic and common

The PT test is a great means of monitoring oral anticoagulant therapy because?

It is sensitive to the vitamin K-dependent factors of the extrinsic and common pathways

(Factors VII, II, and X)

The normal range for PT is approximately?

11.0-13.4 sec

Each laboratory must establish their own PT reference ranges based on?

Type of analyzer, reagent, and patient population

The PT normal range varies depending on?

Type of thromboplastin employed in the testing process and the method of clot detection
The PT is prolonged in individuals with?

Factor deficiency, single or multiple

(Also due to FDPs, heparin, and Hct >55%)

The PT is prolonged in patients with polycythemia as a result of a change in the ratio of anticoagulant to
plasma so the amount of citrate should be?

Decreased.

For coagulation testing, the blood to sodium citrate ratio should always be maintained at?

9:1

PT results may be shortened when the plasma is stored for longer than 4 hours at 4C because of?

Cold activation of factor VII

Coagulation control Level 1 should fall _________ the normal range set forth by the laboratory.

Within

Coagulation control Level II should fall __________ the normal range set forth by the laboratory.

Above

The therapeutic range of warfarin therapy is usually considered to be a PT of?

1 1/2-2 times normal

A large number of drugs can interfere with the action of warfarin and Coumadin in vivo either by?

Potentiating or inhibiting its effect on factors II, VII, IX, and X.

Patients with dangerously elevated PTs caused by Coumadin overdose or liver problems may be given?
Vitamin K to stimulate proper factor production or FFP to directly supply the factor necessary to correct
the abnormal PT.

PT procedure

Reconstitute thromboplastin-CaCl reagent.

Pre-warm small amount of reagent at 37C.

Pipette 4 drops (0.2 mL) thromboplastin into well.

Add 2 drops (0.1 mL) of patient plasma into well.

Hit start on fibrometer.

Perform in duplicate.

Results should be within 1 sec of each other.

What type of thromboplastin reagent did we use in the PT lab in class?

Desiccated rabbit brains

What is the name of the screening test used to evaluate the intrinsic and common pathways or more
precisely to measure all of the plasma coagulation factors with the exception of factors VII and XIII?

Activated Partial Thromboplastin Time (aPTT)

The formation of ___________ occurs at a normal rate only if the factors involved in the intrinsic and
common pathways are present in normal concentrations with normal functionality.

Fibrin

What factors are involved in the intrinsic pathway?

VIII, IX, XI, and XII

What factors are involved in the common pathway?

I, II, V, and X

What factors are involved in the extrinsic pathway?

III and VII

In the aPTT, optimal activation is achieved by the addition of contact activators such as?

Kaolin, celite, micronized silica, and ellagic acid

The aPTT reagent includes a platelet ________________ substitute, which eliminates the test's
sensitivity to platelet number and function?

Phospholipid

The endpoint of the aPTT test is?

The time to clot formation after the addition of calcium chloride (0.02 M)

The aPTT is also used to screen for inhibitors of the intrinsic pathway such as?

Lupus anticoagulant

The aPTT can be used to monitor?

Heparin therapy

The normal range for the aPTT is?

32-37 sec

An inherited or acquired deficiency of factors I, II, V, VIII, IX, and XII, Wafarin or drug interactions, liver
disease, heparin, FDPs, and lupus inhibitor can all cause an aPTT to be?

Prolonged

The PT and aPTT tests measure certain plasma proteins which participate in?

Clot formation

The time for clot formation is detected by?

Completion of an electrical current by a fibrin strand (endpoint)

Neither the PT or the aPTT measure?

Platelets, calcium, or factor XIII

The specimen of choice for the PT and the aPTT is?

Citrated platelet-poor plasma (3.2% sodium citrate-FULL TUBE required)

Sources of error for the PT and aPTT include?

Citrate tube not full (short draw), clotted sample, heparin contamination, traumatic draw (tissue factor),
and hemolyzed sample

aPTT procedure

Incubate 2 drops (0.1 mL) activated partial thromboplastin reagent at 37C for 1 min.

Add 2 drops (0.1 mL) patient plasma.

Incubate at 37C for 2 minutes.

Add 2 drops pre-warmed CaCL.

Hit start on fibrometer.

Duplicate. Results should be within 0.5 sec.

***All pre-warmed when using fibrometer***

The formula to calculate the appropriate citrate volume for patients with hematocrit exceeding 55% is?

C= (1.85 x 10^-3)(100-HCT)(Vol. of blood added)

C is the volume of citrate to remove from tube

Name the medicine used to fight clots that affects the intrinsic pathway.

Heparin

Name the medicine used to fight clots that affects the extrinsic pathway.

Warfarin (AKA Coumadin)

COMPATIBILITY TESTING

What is compatibility testing?

All the steps of ID & testing of donor unit & recipient blood

What is crossmatching?

Mixing of donor RBC & patient serum / plasma

Agglutination / hemolysis: incompatible

What are the required AABB standards of recipient blood?

Positive ID (2) of recipient & their blood sample

Proper sample collection, labeling, handling

ABO & Rh typing recipient blood

Clinically significant antibody testing

Review recipients past blood bank records

What are the required AABB standards of donor blood?

Confirm ABO of donor unit

Confirm Rh if unit is Rh negative

What is required by AABB if recipient is determined to have clinically significant antibody?

Donor units must be phenotyped to prove corresponding antigen is absent

What are the optional AABB standards?

Patient location, sex, diagnosis, date of proposed transfusion

What is considered proper labeling on a blood sample?

Must have 2 independent identifiers, phlebotomist must sign

Sample labeled at bed side

Can't use room # for ID

What must a transfusion request contain?

2 unique identifiers

Component requested & quantity

Name of ordering physician & phlebotomist

Special processing requested

Serum or plasma, testing of which will pick up complement-dependent antibodies?

Serum

If time is critical, test plasma

Why are samples & segments kept at least 7 days post transfusion?

In case of adverse reaction, re-testing may indicate what went wrong

Must recipient blood be tested for weak D?

Nope. If result is D-, patient transfused with D- blood anyways

What must be done before a hospital can use donor blood?

Repeat forward testing of ABO to confirm

D testing only if labeled D-

Testing done on segment attached to the unit

How should matching be done when giving whole blood?

Exact ABO match to recipient phenotype

What indicates incompatibility in crossmatching?

Hemolysis or agglutination at any phase

How much should one unit of transfused RBC increase hematocrit & hemoglobin?
Hct: 3%

Hb: 1g/dL

What does the AABB define as crossmatching?

Methods that demonstrate ABO incompatibility & clin. sig. Ab to red cell Ag & includes AHG test

If no clin. sig. Ab detected, immediate-spin fulfills detection requirement

What is a major crossmatch?

Patient serum & donor red cells

IAT not required if Ab screen is negative

Required if >2mL of RBC in product

What is a minor crossmatch?

Donor serum & recipient red cells

Ab should dilute out anyways when given to patient

Why was it bad for doctors to have blood crossmatched even if blood not typically required?

Once crossmatched, unit is requisitioned & "tied-up"

What should be done if a type & screen shows an atypical antibody?

Antigen negative units should be identified & either reserved or crossmatched

What is done in a type & screen?

Check records

ABO test, Rh type

Do antibody screen

When is an immediate spin crossmatch performed?

Recipient shows no evidence of antibody in record or sample

Detects compatibility, not necessarily exact ABO match

When does AABB require AHG for crossmatching?

Patient demonstrates clin sig Ab in current sample or history

Autoadsorption may be used if autoantibody present

When is an IAT crossmatch required?

Alloantibody, autoantibody, or history of alloantibody

What criteria must be met for computer crossmatching?

FDA criteria of computer system

2 independent determinations of recipient blood type

Confirmation of donor ABO/Rh

Method to verify correct data entry

Discrepancy alert system

Why might a positive crossmatch have a negative antibody screen?

ABO incompatibility

Antibody to low incidence antigen

Positive DAT

What must be on a tag for a crossmatched donor unit?

Patients full name & 2 identifiers

Name of blood product

Unique donor unit number, expiration date, ABO & D type

Crossmatching interpretation

Technologist identification
When can unused blood products be reissued?

Closure hasn't been entered &:

Unit kept within °C conditions, or at room temp if returned within 30 minutes

If a patient has formed a clinically significant antibody, is a compatible cross-match sufficient?

Nope, must phenotype unit with commercial antiserum (verifies antigen negative unit)

What is a massive transfusion?

Total volume of blood is exchanged within 24 hours (10-12 units whole blood)

What is done once ABO identical units are administered in massive transfusion?

Take new recipient sample to check for passively transfused anti-ABO antibodies

What is MSBOS?

Maximum Surgical Blood Ordering Schedule

Average # of blood units allowed to be crossmatched for surgical procedures

Do frozen plasma, PLT concentrates, & cryoprecipitate need to be crossmatched?

Nope

What are the guidelines for selecting platelets?

First choice: ABO & Rh specific

2nd: ABO compatible, Rh specific

3rd: ABO incompatible, Rh specific

4th: ABO & Rh incompatible

HEPATITIS TESTS

What are causes for hepatitis?

virus, ETOH, drugs or toxins, autoimmune

What types of hepatitis are most common?

A, B, & C

If you have hepatitis D what must you also have?

hepatitis B

How does hepatitis damage the liver?

invades hepatocytes and multiples. When the immune system attacks the hepatocytes AST & ALT are
released from damaged cells leading to necrosis and fibrosis.

What happens to albumin and coagulation factors during cirrhosis?

production is decreased and serum albumin will be low and PT will be prolonged

What are the SXS of acute phase hepatitis?

asymptomatic or mild, anorexia, NV, fatigue, HA, malaise, & mild fever, abdominal pain, arthralgia

What are the SXS of chronic phase hepatitis?

pruritus, choluria, jaundice, alcoholic stools

What are the overall signs of hepatitis?

jaundice & scleral icterus, hepatomegaly, splenomegaly (+/-)

acute > 25 X ULN

chronic > 10 X ULN

What type of hepatitis is infectious, has an incubation period of 2-6 weeks, high contagious ad
transmitted by oral-fecal contamination of food and drink?

hepatitis A virus

How is hepatitis A virus identified?

by specific antibodies

What is the outcome of hepatitis A?

complete recovery w/ return to normal LFTs, never becomes chronic, and fulminant hepatic failure < 1%

When is HAV-Ab/IgM seen?

appears 3-4 weeks after exposure and present just before LFTs elevate

When does HAV-Ab/IgM return to normal?

8 weeks

When is HAV-Ab/IgG seen?

appears in 4 weeks after IgM begins to rise

-can remain detectable for years after infection

If IgM ab is elevated in absence of IgG Ab what type of infection is suspected?

acute

If IgG is elevated in absence of IgM elevation what type of infection is suspected?

convalescent stage (recovering from sickness or partially recovered)

What type of hepatitis has an incubation period of 5w-6m (avg 2mo), transmitted via contact w/
infected blood, semen or other bodily fluid and crosses the placenta to newborns of infected moms?

hepatitis B
What type of hepatitis can cause severe hepatitis leading to liver failure and death and has a 10%
change of developing into a chronic state?

hepatitis B

What are complications of hepatitis B?

cirrhosis & primary hepatocellular CA

What type of antigen is the first to indicate infection, rises before clinical symptoms, and indicates active
HBV infection (acute or chronic)

Hepatitis B Surface antigen (HBsAg)

When is HBsAg considered chronic?

persists > 6 months

What type of antibody appears after disappearance of the surface antigen, signifies end of acute phase
and usually persists for life?

Hepatitis B Surface antibody (HBsAb)

What denotes immunity after HBV vaccine or HBV infection?

HBsAb

When are concentrated forms of HBsAb given?

to those who have been exposed to HBV infected pts

What is made by a pts body in response to hep B core antigen?

hepatitis B core antibody (HBcAb)

-expressed but not detectable in serum & not measured

When is HBcAb elevated?

during time lab between disappearance of HBsAg and appearance of HBsAb

"core window"

What 3 things can HBcAB detect?

acute infection, convalescent period, and chronic hepatitis B infection

What 2 types does HBcAb consist of?

IgM anti HBcAG (new infection)

IgG anti HBcAH (old infection)

Describe hepatitis B-e antigen (HBeAg)

protein from the hep B virus that circulates in infected blood when the virus is actively replicating

What antigen is used as an index of infectivity and not for diagnosis, signifies early active disease w/ high
infectivity and will remain elevated (indicating chronic HBV infection)

hepatitis B-e antigen (HBeAg)

What does resolution after a positive test for HBeAg mean?

Hep B has resolved

What antibody indicates that the acute phase of HBV is over and infectivity is greatly reduced in a
chronic condition?

Hepatitis B-e antibody (HBeAb)

What is a direct measurement of the HBV viral load and used serially to monitor response to therapy?

Hepatitis B DNA

How long do you treat HIV?

until levels are undetectable X 2

How can you tell if viral resistance and therapy are not working?

1-2 log increase

100,00-1 billion IU/mL (high rate of HBV replication)

What type of hepatitis has the same route of transmission as Hep B, an incubation period of 2-12 weeks
(avg 6-8) and becomes chronic in 60% of infected pts?

Hepatitis C

AKA non A and non B

How many HCV pts develop cirrhosis?

20%

-increased risk for hepatocellular cancers

When can anti-HCV antibodies be detected?

within 2-6 mo after exposure via EIA

When do anti-HCV antibodies drop below detection limits?

after infection has been cleared

What is HCV-RNA used?

direct detection, monitor dz, and follow response to therapy

When is an acute infection of Hep C determined?

recent exposure and +/- symptoms

If a pt has a pos HCV RNA and a neg anti-HCV antibodies what does the person have?
acute Hep C infection

If a pt has a pos HCV RNA and anti-HCV antibodies w/ documentation of negatives tests w/in the prior 6
months what is it suggestive of?

acute HCV

How do you know when hep C has been cleared?

HCV RNA test neg @ 6 mon and anti-HCV antibodies may remain positive for years

How do you know when a person has a chronic infection of Hep C?

high persistent, non fluctuating HCV RNA levels and positive anti-HCV antibodies

How many people have HVD because of HBV?

5%

Why is Hep D considered a superinfection?

HDV infects a pt who already has chronic HBV

How is HDV transmitted?

via tainted blood

-especially prevalent in drug users and dialysis pts since same mode of transmission as HBV

What is the first parameter to rise (1-3days_, indicates active HDV infection, and is short lived and may
be missed?

HDV-Ag

What is a direct measure of the virus RNA?

HDV RNA

What rises in about 10 days and indicates active HDV infection?

HDV-Ab/IgM

What rises in about 2-3 months and indicates chronic infection?

HDV-Ab total (Igm+IgG)

DENGUE TESTING

Dengue

a mosquito borne disease is a major public

health problem throughout the tropical world.

Incubation period

1-4 days/ 3-14 days

Probable Dengue fever, Dengue with warning signs, and Severe Dengue presenting a:

○ Severe plasma leakage

○ Severe hemorrhage

○ Severe organ impairment

Confirmatory test

PCR

Dengue Fever is confirmed by requesting:

Viral culture and PCR

Probable Dengue fever

Lives in or travels to dengue-endemic area, plus patient has fever and 2 of the following:

Headache

Bod malaise

Myalgia

Arthralgia

Retro orbital pain

Nausea and vomiting

Anorexia

Diarrhea
Flushed skin

Rash

Tourniquet test positive

Leukopenia

Thrombocytopenia

Dengue NS1 antigen test

Dengue with warning signs

Lives or travels to dengue endemic areas

Fever

Abdominal pain

Mucosal bleeding (epistaxis or gum bleeding)

Lethargy

Liver enlargement

Fluid accumulation

Decrease or no urine output

Persistent vomiting

Increased in HCT and thrombocytopenia

Severe Dengue

Lives or travels to dengue endemic areas

Fever

Severe plasma leakage (fluid accumulation with respiratory distress)

Severe organ impairment

Severe bleeding

3 phases of dengue

Febrile
Critical

Recovery

Febrile phase

Fever for 0 to 7 days

Biphasic fever pattern

If the patient is having persistent vomiting and abdominal pain consider these as worrisome findings

Critical phase

Monitor defervescence and warning signs

rapid decrease in platelet count with associated rise in hematocrit and presence of warning signs

Clinical problems in febrile phase

○ Dehydration

○ Hyponatremia

○ febrile seizures

○ neurologic disturbance

Clinical problems in critical phase

○ hypovolemic shock due to plasma leakage

○ organ impairment from prolonged shock

Recovery phase

Gradual reabsorption of extravascular compartment fluid in 48-72 hours

Agent

☛4 serotypes= DENV 1,2,3,4

☛flavivirius genus
☛All serotypes ass.with epidemics of dengue fever (w/ wo DHF) with varying degree of severity.

☛4 serotypes antigenically similar

☛but different enough to elicit cross protection for a few months after infection

☛Secondary infection with dengue serotype 2 or multiple infection with different serotypes= Severe
DHF/DSS

What is the pathogenesis of severe syndrome?

Second infection ➽ virus antibodies are formed within a few days ➽non-neutralizing enhancing Ab
promote infection of higher numbers of MNC ➽release of cytokines, vasoactive mediators, and
procoagulants ➽DIC

Reservoir of Infection

Man and Mosquito

VECTOR

☛Aedes aegypti

☛Aedes albopictus

carry high vectorial competency for dengue virus, i.e., high susceptibility to infecting virus, ability to
replicate the virus and ability to transmit the virus to another host.

Aedes aegypti

☛highly domesticated

anthropophilic

☛nervous feeder (bites more than one host to complete one meal)

☛discordant species (needs more than one feed for the completion of the gonotropic cycle)

☛So, multiple cases and the clustering of dengue cases in the cities

☛breeds in containers, water based air coolers, tyre dumps

Aedes albopictus
☛aggressive feeder

(complete its blood meal in one go on one person)

☛concordant species cities

(does not require a second blood meal for the completion of the gonotropic cycle)

☛*invades peripheral areas of urban cities

Transmission

☛becomes infective by feeding on a patient from the day before onset to the 5th day (viraemia stage) of
illness.

☛After an extrinsic incubation period of 8 to 10 days, the mosquito becomes infective, and is able to
transmit the infection.

☛Once the mosquito becomes infective, it remains so for life.

☛ Genital tract of the mosquito gets infected and transovarian transmission of dengue virus occurs
when virus enters fully developed eggs at the time of oviposition.

Lab findings

☛Leucopenia

☛Neutropenia

☛Thrombocytopenia

☛Elevated ALT or AST

C-REACTIVE PROTEIN

What is C-reactive protein?

Acute phase protein

Released in inflammation & tissue damage

Disappears when inflammation subsides

Lacks disease specificity

Elevated in many disorder

What disorders have elevated levels of C-reactive protein?

Bacterial and viral infections

Rheumatic fever

Malignant diseases

Rheumatoid arthritis

Tuberculosis

Post-op

Myocardial infarction

What are the characteristics of C-reactive protein?

Appears rapidly after acute tissue injury

4 - 6 hours

Concentration may rise to 1000X normal

Reference range: <0.5 mg/dL

Heat labile

Destroyed at 70EC for 30 minutes

Synthesized by liver

What is the role of C-reactive protein in opsonization?

Binds (calcium dependent) to C-polysaccharide in bacterial cell walls

Complement activation
Innate immunity

What is High sensitivity CRP (hs-CRP)?

Used to assess risk for acute coronary syndrome

Stimulated by IL-6 during inflammation

Increases are minimal

Often less than reference range for conventional CRP

What does elevated hs-CRP indicate?

Baseline elevations indicate higher risk for coronary artery disease and death from CAD with or without
presence of clinical symptoms.

What do treatments to decrease inflammation may also reduce?

Aspirin or other anti-inflammatory drugs reduce risk of coronary artery disease

What is does it mean to have 0-1 mg/L of hs-CRP?

Low risk for acute coronary syndrome

What is does it mean to have 1-3 mg/L of hs-CRP?

Moderate risk for acute coronary syndrome

What is does it mean to have anything over 3mg/L of hs-CRP?

High risk for acute coronary syndrome.

What must be performed with hs-CRP to detect minimal changes?

High sensitivity methods

What are some methods for conventional laboratory detection of CRP?

Agglutination (latex)
Anti-human CRP attached to latex particle

Fluorescent antibody

Precipitation (capillary or tube)

Gel diffusion

Immunonephelometry

RIA

What are the characteristics of CRP assays?

Reference range: < 0.5 mg/dL

sensitive to 3mg/L

Uses

Monitoring inflammation

Evaluating treatment

What are some methods for laboratory detection of hs-CRP?

Immunonephelometry

Uses monoclonal antibody to CRP

Sandwich immunoassay

Sensitive to 0.175 mg/L