Lab Manual GUS

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Genitourinary

System Laboratory Manual

3rd Year Undergraduate Program

Faculty of Medicine Universitas Padjadjaran

3/20/2015



2 Genitourinary System – 3rd Year Undergraduate Program

2014 - 2015

TABLE OF CONTENTS

WEEK DEPARTMENT TOPIC PAGE I Embryology Embryology of GUS 3

II Anatomy Anatomy of GUS 11

III Histology Histology of GUS 16

IV Pathology

Anatomy

Pathology anatomy of

GUS 39

V Physiology Renal Regulation of

Fluid Balance 58

VI Clinical

Pathology

Urine Microscopic

Examination 62

VII Microbiology Microbiology of GUS 68

VIII Pharmacology Drugs Acting on the

kidney; comparison

between some diuretics

agents 79




2014 - 2015

WEEK I

EMBRYOLOGY OF GENITOURINARY SYSTEM

LEARNING OBJECTIVES

After performing laboratory activity, the student should be able to:

1. Describe development of kidney system.

2. Describe molecular regulation of kidney development.

3. Describe abnormalities/anomalies of the kidney development.

4. Describe the development and abnormalities of ureter, urinary bladder, urethra and

prostate.5. Describe development of the indefferent embryo.

6. Describe primordium of the external genitalia.

7. Describe clinical considerations of the development of external genitalia.

RESOURCE PERSON

1. Retno Ekowati, dr., M.Kes

REFERENCES

1. Sadler, T.W. Langman’s Medical Embryology . 10th Ed. Lippincott Williams and

Wilkins. Philadelphia. 2006. pp. 229-256.

2. Moore, K.L., and Persaud, T.V.N. The Developing Human. Clinically Oriented

Embryology . 7th Ed. Elsevier Saunders. 2003. pp. 255-285.




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HOMEWORK ASSIGNMENTS

Complete the figure by naming the pointed parts:

Fig. 2.1 Transverse sections through embryos at various stages of development showing

formation of nephric tubules. A. 21 days. B. 25 days. Note formation of external and internal

glomeruli and the open connection between the intraembryonic cavity and the nephric tubule.

Fig. 2.2 A. Relationship of the intermediate mesoderm of the pronephric, mesonephric, and

metanephric systems. In cervical and upper thoracic regions, intermediate mesoderm is

segmented; in lower thoracic, lumbar, and sacral regions it forms a solid, unsegmented mass

of tissue, the nephrogenic cord. Note the longitudinal collecting duct, formed initially by the

pronephros but later by the mesonephros. B. Excretory tubules of the pronephric and

mesone hric s stems in a 5-week embr o.




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Fig. 2.3 A. Transverse section through the urogenital ridge in the lower thoracic region of a 5-

week embryo showing formation of an excretory tubule of the mesonephric system. Note

the appearance of Bowman’s capsule and the gonadal ridge. The mesonephros and gonad

are attached to the posterior abdominal wall by a broad urogenital mesentery. B. Relation of

the gonad and the mesonephros. Note the size of the mesonephros. The mesonephric duct(wolffian duct) runs along the lateral side of the mesonephros.




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LABORATORY ACTIVITY

1. Discuss homework materials in small group, and explain development of the kidney.

2. Explain molecular regulation of the kidney development.

3. Explain clinical correlation that appear, if any, in the development of kidney.

4. Describe the Anomalies of Kidney.

5. Describe: The development of Ureter and Anomalies of the Ureter.

6. Describe: The development of Urinary bladder and Anomalies of Urinary Bladder)

7. Describe: The development of Male Urethra and Anomalies of Urethra

8. Describe: The development of the Prostate

9.

Describe: The development of Male Genital Externa and Anomalies of Male ExternalGenitalia

10. Describe: The development of Testis and Descent of Testis

Fig. 2.4 Relation of the hindgut and cloaca at the end of the fifth week.

The ureteric bud penetrates the metanephric mesoderm (blastema).

Fig. 2.5 A-C Ascent of the kidney. Note the change in position between the

mesonephric and metanephric systems. The mesonephric system degenerates

almost entirely, and only a few remnants persist in close contact with thr gonad.

In both male and female embryos, the gonads descend from their original level to

a much lower position.




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Complete the figure by naming the pointed parts:

Fig. 2.6 Development of the renal pelvis, calyces, and collecting tubules of the

metanephros. A. 6 weeks, B. At the end of the sixth week, C. 7 weeks, D. Newborn. Note

the pyramid form of the collecting tubules entering the minor calyx.

Fig. 2.7 Development of a metanephric excretory unit. Arrows, the place where the excretory unit

(blue) estblishes an open communication with the collecting system ( yellow ), allowing flow of urine

from the glomerulus into the collecting ducts.




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Fig. 2.8 Genes involved in differentiation of the kidney. A. WT1, expressed by the mesenchyme, enables

this tissue to respond to induction by the ureteric bud. GDNF and HGF, also produced by mesenchyme,

interact through their receptors, RET and MET, respectively, in the ureteric bud epithelium, to stimulate

growth of the bud and maintain the interactions. The growth factors FGF2 and BMP7 stimulate

proliferation of the mesenchyme and maintain WT1 expression. B. PAX2 and WNT4, produced by the

ureteric bud, cause the mesenchyme to epithelialize in preparation for excretory tubule differentiation.

Laminin and type IV collagen form a basement membrane for the epithelial cells .




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Fig. 2.9 A, B. A complete and a partial double ureter. C. Possible sites of

ectopic ureteral openings in the vagina, urethra, and vestibule. D, E.

Photomicrographs of complete and partial duplications of the ureters (U).

Arrows, duplicated hilum; B, bladder; K , kidneys; ML, median umbilical

ligament.

Fig. 2.10 A. Unilateral pelvic kidney showing the

position of the adrenal gland on the affected side. B, C.

Drawing and photomicrograph, respectively, of

horseshoe kidneys showing the position of the inferior

mesenteric artery. BW , bladder wall; U, ureters.




2014 - 2015

----




2014 - 2015

WEEK II

ANATOMY OF GENITOURINARY SYSTEM

LEARNING OBJECTIVES

1. Describe the anatomic topography of kidney.

2. Describe the relationship of kidney and suprarenal gland to adipose, fascial

coverings, and other abdominal organs.

3. Describe the vascularization, innervations and lymphatic vessels of kidney.

4. Describe basic structure gross anatomy of kidney.

5. Describe the branches of abdominal aorta and tributaries of inferior vena cava (level

of vertebrae).6. Describe the anatomic topography of pelvic urinary organs.

7. Describe the anatomical aspects of ureters, urinary bladder and urethra.

8. Describe the vascularization, innervations and lymphatic vessels of ureters, urinary

bladder and urethra.

9. Describe the structure and function of ureters, urinary bladder, and urethra.

10. Compare the urethras of males and females.

11. Describe the anatomic topography of male and female genital organs.

12. Describe the anatomical aspects of male and female genital organs.

13. Describe the vascularization, innervations and lymphatic vessels of male and female

genital organs.

SEQUENCE

1. Pretest 15 minutes

2. Lab activity 120 minutes

a. Video guide

b. Specimen identification

c. Anatomy pictionary quiz

3. Post test 15 minutes

RESOURCE PERSON

1. Putri Halleyana A. R., dr

2. Gita Tiara D. N., dr

3. Nani M. Yazid., drg., Mkes

REFERENCES

1. Moore, KL and Dalley, AF. Clinical Oriented Anatomy. 5th Ed. Lippincott Williams &

Wilkins. 2006. pp. 308-320.




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NOTE

Before starting the lab activity, the student must:

1. Wear the lab coat properly

2. Bring the reference book or atlas

3.

Finish their homework assignment

INTRODUCTION

Functionally, the urogenital system can be divided into two entirely different

components: the urinary system and the genital system.

The urinary system consists of two kidneys, two ureters, one urinary bladder, and

one urethrae. The superior urinary organs (kidneys and ureters) and their vessels are

primary retroperitoneal structures on the posterior abdominal wall.

The kidneys do the major work of the urinary system. They produce urine that is

conveyed by the ureters to the urinary bladder in the pelvis.

HOMEWORK ASSIGNMENTS

1. Please make a drawing of urinary system consist of both kidneys (and it’s attachment

to the IVC and abdominal aorta), both ureters, urinary bladder, and the urethra!

2. Describe the kidney’s:

a. Topography

b. Vascularization. Make a schematic drawing of kidney’s arteries & veins

c. Innervation

3. Describe the ureter’s anatomical course. Including the structures it crosses and

consctrictions

4. Describe the difference of the right and left suprarenal gland

5. Describe the urinary bladder’s:

a. Surface and borders

b. The difference of organ relations in male and female

c. Vascularization

d. Innervation

6. Compare the male and female urethra in a table diagram!

7. Make a drawing of the internal and external male genitalia! Explain the functions of

the organ briefly in the drawing!

8.

Describe the topography, organ relations, vascularization, and innervation of:

a. Female internal genitalia organ

b. Female external genitalia organ




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LABORATORY ACTIVITY

I. Video Introduction

II. Identification

Structure check comment

Urinary System

Kidney

Location

Superior & inferior pole

Topography: right & left kidney

Coverings

Fibrous capsule

Perinephric fat

Renal fascia

Paranephric fat

Surfaces: anterior & posterior

Internal structures

Renal Pyramid

Renal Column

Renal Papilla

Renal Pelvis

Major calices

Minor calicesVascularization

Renal artery & its branching: right & left

difference

Renal veins & its tributaries: right & left

difference

Lymphatic

Innervation

Ureter

Location & course: abdominal part, pelvic part

ConstrictionsVascularization

Arterial

Veins

Lymphatic

innervation

Urinary Bladder

Shape & topography: empty vs filled, male vs

female

Fixation

External structure: apex, base, body, neckSurfaces: superior, posterior, infero-lateral




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Internal structure: ureter orifices, urethral orifice,

trigone, detrussor muscle

Vascularization

Innervation

Urethra

Male: prostatic part, intermediate part, spongy part

Female: course

Sphincter

Vascularization

Innervation

Genitalia

Male

Internal: topography, course, vascularization,

innervation

Testes

Epididymides

Ductus deferentes

Seminal vesicles

Ejaculatory ducts

Prostate: topography, organs relation, parts

surfaces, ducts

Bulbourethral gland

Spermatic cord

External

Scrotum

Penis

External structure

Internal structure

Muscles & ligaments

Vascularization

Innervation

Female

Internal: topography, course, vascularization,

innervationVagina

Uterus

Uterine tubes

Ovaries

External: topography, course, vascularization,

innervation

Mons pubis

Labia majora

Labia minora

ClitorisBulbs of vestibule




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Greater vestibular glands

III. Anatomy pictionary quiz

Each group will be ask to hand 20 pictionary questions to be guessed by the other

group. The number of guessed question will determine the winner and loser. The

winning group will be given a privilege while the losing group will be given extra

anatomy assignment.

Trainer’s authorization




2014 - 2015

WEEK III

HISTOLOGY OF GENITOURINARY SYSTEM

LEARNING OBJECTIVES

After performing laboratory activity, the student should be able to:

1. List the organs of the urinary system and male genital system

2. Identify the structures and regions of a kidney and describe their organization

3. Describe the structure, function, and location of each component of a nephron

4. Describe the function of juxtaglomerular apparatus and identify its component

5. Trace the flow of blood through the kidney

6.

Identify the components of the glomerular filtration barrier in a diagram of a portionof a renal corpuscle

7. Describe the structure and function of ureters

8. Describe the structure and function of urinary bladder

9. Describe the structure of male urethra

10. Describe the location of the glands, ducts, and external genitalia of the male

reproductive system

11. Describe the general organization of the testis

12. Describe the differences in wall structure of excretory genital ducts.

13. Compare the prostate, seminal vesicles, and bulbourethral glands in terms of general

organization, epithelial lining, secretory products, and the point(s) at which theirsecretions enter the excretory pathway.

RESOURCE PERSON

Nursiah Nasution, dr. (Department of Histology)

REFERENCES

1. Junqueira, L.Carlos. Basic Histology Text and Atlas. 12th Edition. Lange Medical Books

McGraw-Hill.

2. Young, B and Heath, J.W.Weather’s Functional Histology. 4th Edition. Churchill

Livingstone.

3. Paulsen, Douglas F. Basic Histology Examination & Board Review . 2nd Edition.

Appleton & Lange.

4. Gartner, Leslie P and Hiatt, James L. Color Textbook of Histology . 2nd Edition. Saunders




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INTRODUCTION

The urinary system includes the kidneys and the urinary tract. Kidneys are paired

bean-shaped, retroperitoneal organs are located in the posterior wall of the abdominal

cavity. The components of urinary tract are ureters, urinary bladder, and urethra. The lumen

of the tract is characteristically lined by transitional epithelium, except for certain portionsof the urethra. The kidneys filter metabolic wastes and foreign substances from the blood;

regulate the ion, salt, and water concentrations of the fluids that bathe the body’s tissues;

and produce rennin and erythropoietin. The collection of raw filtrate from the blood in the

glomerular capillaries is only the first step in urine production. It is followed by the

reabsorption of important ions, small proteins, nutrients, and much of the water. These are

returned to the blood in the peritubular capillaries and vasa recta in precise proportions.

The portion of the raw filtrate that is not reabsorbed constitutes the urine; it is carried by

the ureters from the kidneys to the urinary bladder, where it is temporarily stored and later

released through the urethra.

The male genital system consists of the external genitalia and a series of glands andducts that produce and transport the male gametes (spermatozoa) and the seminal fluid.

During laboratory activity, the students will be asked to show any anatomical parts

using model, interactive CD, as well as histological preparations and finally also discuss

about important aspects of physiology and pathogenesis. Before activity, the students have

to accomplish the homework assignment first, read the primary references, so that they will

be more readily performing laboratory activity

The bladder and the urinary passages (calyces and renal pelvis) store urine formed in

the kidneys and conduct it to exterior. The ureters pass through the wall of the bladder

obliquely, forming a valve that prevents the backflow of urine. The urethra is a tube that

carries the urine from the bladder to the exterior. In men, sperm also pass through it duringejaculation. In women, the urethra is exclusively a urinary organ.

The male reproductive system may be divided into four major functional components:

The testes or male gonads, paired organs lying in the scrotal sac, are responsible for

population of the male gametes, spermatozoa, and secretion of male sex hormones,

principally testosterone.

A paired system of ducts, each consisting of ductuli efferentes, epididymis, ductus

deferens and ejaculatory duct, collect, store and conduct spermatozoa from each testis. The

ejaculatory ducts converge on the urethra from which spermatozoa are expelled into the

female reproductive tract during copulation.

Two exocrine glands, the paired seminal vesicle and the single prostate gland, secrete a

nutritive and lubricating fluid medium called seminal fluid in which spermatozoa are

conveyed to the female reproductive tract. Semen, the fluid expelled during ejaculation,

consists of seminal fluid and spermatozoa, plus some desquamated duct lining cells.

The penis is the organ of copulation. A pair of small accessory glands, bulbourethral

glands of Cowper, secreted a fluid which lubricated the urethra for passage of semen during

ejaculation

The accessory genital glands produce secretions that are essential for the reproductive

function in men.

The prostate is a collection of 30-50 branched tubuloalveolar glands. The prostate has 3

distinct zones. The transition zone is of medical importance because it is the site where

most benign prostatic hyperplasia originates




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The seminal vesicles consist of 2 highly tortuous tubes about 15 cm in length they are glands

produce a viscid, yellowish secretion that contains spermatozoa-activating substances.

The bulbourethral glands, is 3-5 mm in diameter. The secreted mucus is clear and acts as a

lubricant.

HOMEWORK ASSIGNMENT

Please complete this figure by naming the pointed parts!

KIDNEY

Pyelum, ureter, renal artery, medullary pyramid, capsule, cortex, renal vein, calyx mayor, calyx

minor

General organization of the kidney (frontal section)




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Please complete this figure by naming the pointed parts of renal vasculature!

Figure: Basic organization of the nephron, collecting system and renal vasculature

Glomerulus, afferent arteriole, efferent arteriole, cortical radial artery. Interlobar artery, arcuate

artery




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Collecting duct, proximal convoluted tubule, distal convoluted tubule, Henle’s loop




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JUXTAGLOMERULAR APPARATUS

Glomerulus, Bowman’s space, Bowman’s capsule, proximal convoluted tubule, distal convoluted

tubule, afferent arteriole, efferent arteriole , macula densa juxtaglomerular cell, lacis cells

Diagram: Juxtaglomerular apparatus




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Podocyte cell body, filtration slit, Primary process, secondary process (pedicel)

Schematic representation of a glomerular capillary with the visceral layer

of Bowman’s capsule (formed by podocytes)




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Please list components of the wall structure of this figure!

Figure No.1: Transverse section of the ureter, Masson’s trichrome

Please list components of the wall structure of this figure!

Figure No.2: Section of wall layer the urinary bladder, Masson’s trichrome




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60

Drawing of a transverse section of the penisDrawing of a transverse section of the penis

Corpuscavernosumof the penis

Erectile

tissue

Corpuscavernosumof the urethra

Urethra

Dorsalarteries

Superficialdorsal vein

Deep dorsalvein

Tunicaalbuginea

Deep artery

Figure No. 3

Urethra, corpus cavernosum of the urethra, corpus cavernosum of the penis, dorsal arteries,

superficial dorsal vein, deep artery, deep dorsal vein, tunica albuginea, erectile tissue




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Testis

testicular lobules, seminiferous tubules, septae, tunica albuginea, rete testis, tunica vaginalis

(parietal layer), tunica vaginalis (visceral layer), serous space, ductuli efferentes, ductus

deferens, epdidiymis

Figure: testis




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44

SEMINIFEROUS TUBULES, STRAIGHT TUBULES,

RETE TESTIS AND DUCTULI EFFERENTES

1. Seminiferoustubules

2. Straighttubules

3. Connectivetissue ofmediastinum

4. Rete testistubules

5. Ductuliefferentes(efferentductules

6. Rete testistubules

Rete testis tubules, ductull efferentes, rete testis tubule, connective tissue of mediatinum,

seminiferous tubules, straight tubules




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46

1. Connectivetissue

2. Cross sectionsof the ductusepididymidis

3. Basement

membrane

4. Pseudostratifiedcolumnarepithelium withstereocilia

5. Sectionthrough of U-bend of theductusepididymidis

6. Epididymalwall cuttangentially

7. Smoothmuscle fibers

8. Stereocilia

9. Columnar cells

10. Basal cell

Ductus Epididymidis (Duct of the Epididymis)Ductus Epididymidis (Duct of the Epididymis)

Basement membrane, connective tissue, cross sections of the ductus epididymidis,

pseudostratified columnar epithelium with stereocilia, basal cell, columnar cells,

epididymidis wall cut tangentially, stereocilia, section through of U-bend of the ductus

epididymidis




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Please complete this figure by naming the pointed parts

49

1. Outerlongitudinalmuscle layer

2. Circularmuscle layer

3. Innerlongitudinalmuscle layer

4. Nerve andblood vesselsin theadventitious

5. Lamina propria

6. Longitudinal crestof lamina propria

7. Epithelium

8. Adipose tissue

Ductus Deferens (Transverse section)Ductus Deferens (Transverse section)

Lamina propria, epithelium, adipose tissue, longitudinal crest of lamina propria, inner

longitudinal muscle layer, outer longitudinal muscle layer, circular muscle layer, nerve and

blood vessels in the adventitious




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Central zone, peripheral zone, transition zone


57

4. Glandularepithelium

5. Ducts

6. Smooth

muscle fibers

7. Capillary andvenule

1. Prostaticconcretions

2. Glandularalveoli

3. Glandularducts

Prostate GlandProstate Gland

Glandular ducts, glandular alveoli, glandular epithelium, prostatic concretions, smoothmuscle fibers, capillary and venule.




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57

Prostate Gland with Prostatic UrethraProstate Gland with Prostatic Urethra

1. Prostaticurethra

2. Colliculusseminalis

3. Fibromuscular

stroma

4. Prostaticglands (alveoli)

5. Prostaticglands (alveoli)

6. Smooth muscle ofthe stroma

7. Dilatation ofthe utriculus

8. Diverticula ofurethral wall

9. Utriculus

10. Ejaculatoryducts

Fibromuscular stroma, colliculus seminalis, prostatic glands (alveoli), prostatic urethra,

utriculus, ejaculatory ducts, diverticula of urethral wall, dilation of the utriculus, smooth

muscle of the stroma




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52

Seminal VesicleSeminal Vesicle

4. Glandularepithelium

5. Primary fold inthe mucosa

6. Secondary folds

7. Lamina propria

1. Crypts in themucosa

2. Muscular coat

3. Adventitia

Lamina propria, primary fold in the mucosa, secondary folds, glandular

epithelium, crypts in the mucosa, adventitia, muscularis coat

Think first and answer the question correctly and concisely!

Identify the structures and regions of a kidney and describe their organization

1. List the organs of urinary system and male genitale system

2. Describe the structure, function, and location of each component of a nephron

3. Describe the function of juxtaglomerular apparatus and identify its components

4. Trace the flow of blood through the kidney

5.

Identify the components of the glomerular filtration barrier in a diagram of a portionof a renal corpuscle

6. Describe the structure and function of ureters

7. Describe the structure and function of urinary bladder

8. Describe the structure of male urethra

9. Describe the location of the glands, ducts, and external genitalia of the male

reproductive system

10. Describe the general organization of the testis

11. Describe the differences in wall structure of excretory genital ducts.

12. Describe the microstructure of prostate gland

13.

Describe the microstructure of seminal vesicles14. Describe the microstructure of bulbourethral glands




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LABORATORY ACTIVITY

Pre-requisites: The students have to do the homework assignment and read the references

as listed in the first page.

Task:

1. Discus the homework materials in the small group (tutorial group).

2. See the specimen under microscope and try to make a schematic draw and put the

most important description based on the schematic draw.

Specimen No.1 (kidney, H.E.)

Schematic Draw ( cortex) Description




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Specimen No.2 (kidney, H.E.)

Schematic Draw (medulla) Description

Specimen No.3: ureter

Schematic Draw Description




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Specimen No.4: urinary bladder


Specimen No.4: transverse section of the penis





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Specimen No.5: Seminiferous tubules


Specimen No.6: Tubuli recti and rete testis





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Specimen No.7: Ductuli efferentes


Specimen No.8: Ductus epididymidis





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Specimen No.9: Ductus deferens


Specimen No.10: prostate gland





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Specimen No.11: prostate gland with prostatic urethra


Specimen No.12: seminal vesicles





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WEEK IV

PATHOLOGY ANATOMY OF GENITOURINARY

SYSTEM

LEARNING OBJECTIVES

After performing laboratory activity, the students should be able to :

1. Explain the macroscopic and microscopic of post streptococcal glomerulonephritis

2. Explain the macroscopic and microscopicof chronic pyelonephritis.

3. Explain the macroscopic and microscopicof common kidney tumour

4. Explain the macroscopic and microscopicof polycystic kidney

5. Explain the macroscopic and microscopicy ofcystitis

6.

Explain the macroscopic and microscopicof urothelial carcinoma of the bladder

7. Explain the macroscopic and microscopicof benign prostatic hyperplasia

8. Explain the macroscopic and microscopicof prostatic adenocarcinoma

9. Explain the macroscopic and microscopicof seminoma of the testes

RESOURCE PERSON

1. Hasrayati Agustina, dr.,SpPA

2. Hermin Aminah, dr,. SpPA

REFERENCE

Kumar V, Abbas AK and Fausto N. Robbins and Cotran Pathologic Basis of Diseases ( 2005),

7th Edition. Elsevier Saunders. pp .504-506, 960-1019

HOMEWORK ASSIGNMENT

A. Fill in the blank box under each the picture with microscopic and macroscopic

appearance of each cases

LABORATORY ACTIVITY

Pre-requisites: The students have to do the homework assignment and read the references

as listed in the first page.

Task:

1. Discuss the homework materials in the small group (tutorial group).

2. See the specimen under microscope and try to make a schematic draw and put the

most important description based on the schematic draw.




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1. ACUTE POST STREPTOCOCCAL GLOMERULONEPHRITIS

A 10-year-old boy had just recovered from skin infection ( impetigo ).Two weeks

later, he developed malaise, nausea and slight fever , he passed dark brown urine.

Microscopic :




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1 = 2 = 3=

12

3

4

5




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2. CHRONIC PYELONEPHRITIS

A 40-year-old man presentedto the hospital because of recurrent urinary tract

infections. He complained back pain, fever, polyuria and his blood pressure was 170/100

mmHg. A renal sonogram revealed extensive scarring with pelvic and calyceal enlargement

and also cortical thinning.

Macroscopic :

Microscopic :




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3. WILM’S TUMOR

A 3-year-old boy presented to the hospital because of a large abdominal mass. He

had hematuria, abdominal pain and constipation. USG examination revealed a large mass in

his left kidney.

Macroscopic :

Microscopic :




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4. RENAL CELL CARCINOMA

A 60-year-old man presented to the hospital because of flank pain and hematuria

since 1 month ago. He also complained fever, weakness and weight loss.

Macroscopic :

Microscopic :




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5. POLYCYSTIC KIDNEY

A 5 years old girl presented to the hospital because of recurrent urinary tract infection. USG

reveal cystic masses in both of her kidney

Macroscopic

Microscopic




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Schematic draw

Specimen No 2: Specimen No 3:

Specimen No 4 : Specimen No 5 :




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6. CHRONIC NON SPECIFIC CYSTITIS AND CYSTITIS CYSTICA

A 37 years old female came to the outpatient clinic because of disuria and frequency since 2 weeks ago .She

had no fever or flank pain

Macroscopic




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7. UROTHELIAL CARCINOMA OF THE BLADDER

A 56-year-old male had several episodes of hematuria and dysuria in the past few week. The patient

was a chronic smoker for 30 years. A urinalysis show 4 (+) blood. The urine culture is negative. A cystoscopy

was performed, and a 2 cm, sessile, friable mass was seen on the left lateral wall.

Microscopic:

Macroscopic:




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8.

BENIGN PROSTATIC HYPERPLASIAA 65-year-old male had increasing difficulty in urination. He had to get up several

times at night because of a feeling of urgency. He has difficulty starting and stopping

urination. His serum PSA level was slighty increased.

Microscopic :

Macroscopic:




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9. PROSTATIC ADENOCARCINOMAA 65-year-old male presented with a long history of urinary hesitancy nocturia.

Digital rectal examination revealed a hard, irregular prostate. Serum PSA level is 120 mg/ml.

Microscopic :

Macroscopic :




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Microscopic:




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Specimen No 1:





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Specimen No 2:


Specimen No 3 :





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Specimen no 4 :





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10. SEMINOMA OF THE TESTES

A 57 year-old male had a unilateral enlargement of the testss. Physical exanination

showed enlargement of the left testes 5 cm in diameter

Macroscopic :




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Microscopic :




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Specimen No 1:





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WEEK V

PHYSIOLOGY OF GENITOURINARY SYSTEM:

RENAL REGULATION OF F LUID B ALANCE

LEARNING OBJECTIVES

After performing laboratory activity, the students should be able to:

1. Describe the renal regulation of fluid balance.

2.

Demonstrate and explain how the kidneys respond to the water loading and salt waterloading by analyzing changes in urinary volume, specific gravity and pH.

RESOURCE PERSON

1. Dr. med. Setiawan, dr. (Physiology)

2. Reni Farenia, dr., M.Kes (Physiology)

3. Yuni Lazuardi, dr., M.Kes (Physiology)

4. Nova Sylviana dr.M.Kes ( Physiology)

REFERENCES

1. Guyton AC and Hall JE. Textbook of Medical Physiology. 11th Edition. Elsevier

Saunders. pp 348-365.

INTRODUCTION

Human being can be fasting of foods for several days. But, this survival capacity is

very limited for water. Exposing to water challenges require the capability of regulatory

process which mostly involves the kidney ability. Urine volume, solute concentration, and

electrolyte content are adjusted by the kidneys to maintain homeostasis of the blood.Drinking excess water or eating salty foods results in a rising blood volume, which is

followed by compensatory increases in the urinary excretion of the salt and water.

Diuretics are agents that increase the rate of urine formation (diuresis). These agents

are ion transport inhibitors that decrease the reabsorption of Na+ at different sites in the

nephron. As a result, Na+ and other ions such as Cl- enter the urine in greater amounts

rather than normal, along with water, which is carried passively to maintain osmotic

equilibrium. The efficacy of the different classis of diuretics vary considerably, with the

increase in secretion of Na+ varying from less than 2% for the weak, potassium-sparing

diuretics, to over 20% for the potent loop diuretics. Their major clinical uses are in managing

disorders involving abnormal fluid retention (edema) or in treating hypertension where




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their diuretic action causes decreased blood volume, which leads to reduction in blood

pressure.

HOMEWORK ASSIGNMENT

1. How is the mechanism of water regulation reflected by urinary excretion once a person

is challenged with drinking excess water for instance in amount of 1 L?

2. What do you expect from the urinary excretion analysis (urinary volume, spesific gravity

and pH) in above condition?

LABORATORY ACTIVITY

Pre-requisites: The students have to do the homework assignment and read the references.

Task of Renal Regulation of Fluid Balance

Materials:

1. Urine collection cups

2. Urinometers

3. pH paper (pH range 3 –9)

Procedure:

1. The students void their urine into collection cups at the beginning of the laboratory

session. In the analyses done in step 4, this sample will serve as thecontrol (time zero).

2. The students drink 1000 mL of water or salt containing water (4 gram of NaCl).

3. After drinking the solutions described in step 2, the students void their urine every 30

minutes for 2 hours. The urine samples are analyzed as described in step 4.

4. Each of the five urine samples collected are analyzed for volume, pH, and specific gravity,

as follows:

(a) Volume (mL). Measure the approximate volume of urine obtained and enter the data in

the table of the laboratory report.

(b) pH. Determine the pH of the urine samples by dipping a strip of pH paper into the urine

and matching the color developed with a colorchart. The urine normally has a pH

between 5.0 and 7.5.

(c) Specific gravity. Determine the specific gravity of the urine samples by floating a

urinometer in a cylinder (fig. 9.3) nearly filled with the specimen. Read the specific

gravity at the meniscus on the urinometer scale, making sure that the urinometer float




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is not touching the bottom or the sides of the cylinder. The specific gravity is directly

related to the amount of solutes in the urine and ranges from 1.010 to 1.025. (Pure

water should have a specific gravity of 1.000.)

Worksheet

Enter your data in the following table of observation:

Drinking of pure water

Time Urine volume Urine pH Urine Specific gravity

0’

30’

60’

90’

120’




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Drinking of salt containing water

Time Urine volume Urine pH Urine Specific gravity

0’

30’

60’

90’

120’

Application of the knowledge:

1. How is the urine analysis results if a person is now challenged with the high salt

containing water?

2. Could you explain the regulatory mechanism involved?

3. What would be happened in a dehydrated person?

---------------




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WEEK VI

CLINICAL PATHOLOGY OF GENITOURINARY

SYSTEM: URINE MICROSCOPIC E XAMINATION

HOMEWORK ASSINGMENT

To be collected to your tutor at the day of lab activity

What will you do on your laboratory activity today?

I. OBJECTIVE

At the end of the activity the students will understand and can describe about:

1. The recommended of preanalyticalstage of urine examination

2. Analytical stage: The urine microscopic examination

3. Post analytical stage of urine examination

4.

The interpretation of the test results5. Factors which can affect or interfere the results

II. REFERENCE:

1. Strassinger SK; Di Lorenzo MS; Urinalysis and Body Fluids; 4 thed; Philadelphia;

F.A. Davis Company, 2001.

III. INTRODUCTION

The results of urine microscopicexamination contains information about many of the

body’s major information can be obtained by inexpensive laboratory tests.

IV. INTENDED USE

Reasons for performing the urinalysis include aiding in the diagnosis of disease,

screening asymptomatic populations for undetected disorders, and monitoring the

progress of disease and the effectiveness of therapy.

V. EQUIPMENTS




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In this laboratory activity the students do the microscopic analysis of the urine by

light microscope. Another equipments: object glass, cover glass, glove, and tissue.

VI. PROCEDURES

PREPARATION/PREANALYTIC PROCEDURE1. Urine pot: Dry, clean, leak-proof (disposable pots are recommended).

2. Midstream urine:

- Patients must be provided with appropriate cleansing materials, sterile

pot and instructions for cleansing and voiding.

- Patients are instructed to wash their hands prior to beginning the

collection. Male patients should clean the glands beginning at the urethra

and withdrawing the foreskin. Female patients should separate the labia

and clean the urinary meatus and surrounding area.

ANALYTICAL PROCEDURE

A. PHYSICAL EXAMINATION

The physical examination of urine includes the determinations of the urine color,

clarity, and specific gravity. The results of the physical portion of urinalysis also

can be used to confirm of or to explain findings in the microscopic examination of

urinalysis.




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TEST PROCEDUREOF PHYSICAL EXAMINATION

Determination of color and clarity can do on urine in a clean, dry and leak-proof container.SG can

also determine with a refractometer.

This procedure must be followed exactly to achieve reliable results. 1. Use your gloves

2. Collect fresh, well-mixed, uncentrifuged urine specimen in a clean dry

container. Mix well immediately before using.

B. MICROSCOPIC EXAMINATION:

The microscopic examination of the urineuse a sediment of urine that had

centrifugated. Its purpose is to detect and to identify insoluble materials present

in the urine. The blood, kidney, lower genitourinary tract, external

contaminations all contribute formed elements to the urine. These include RBCs,

WBCs, epithelial cells, casts, bacteria, yeast, parasites, mucus, spermatozoa,

crystals, and artifacts. Examination of the urinary sediment must include both

identification and quantitation of the elements present.

Squamous epithelial cells




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Squamous epithelial cells

White blood cell and granular castStained white blood cell cast

Fatty cast Mucus

Uric acid crystals Calcium oxalate crystals




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Triple phosphate crystalsCystine crystals

Cholesterol crystals

POSTANALYTICAL: Recording and Reporting

MICROSCOPIC EXAMINATION

- RBCs: <1/hpf

- WBCs: < 6/hpf

- Epithelial cells: represent normal sloughing of old cell. Three types of epithel cells are

seen in urine: squamous, transitional, and renal tubular.

- Casts: negative

- Bacteria, yeast, parasites: negative

-

Mucus: normal :a small amount. It has no clinical significance when present in either

female or male urine

- Spermatozoa: occasionally found in the urine of male

- Crystals: depends on the pH of urine; normal in acid urine: uric acid, acid urates, and

sodium urates; in alkaline urine: amorphous phosphate, triple phosphate, calcium

phosphate, calcium carbonate, and ammonium biurate. In abnormal urine you can

found: cystine, cholesterol, leucine, thyrocine, bilirubin, sulfonamides, ampicillin, and

radiographic dye.




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RESULT:

……………………………………………………………………………………………………………………………………

………………..…………………………………………………………………………………………………………………

………………………….………..………………………………………………………………………………………………

…………………………………………….……..

DISCUSSION:

……………………………………………………………………………………………………………………………..……

……………………………………………………………………………………………………………………………………

……………..……………………………………………………………………………………………………………………

………………………………..……………………................................................................................

..............................................................

CONCLUSION:

……………………………………………………………………………………………………………………………………

……………………………………………………………………………………………………………………………………

……………………………………………………………………………………………………………………………………

……………………………………………………………………………………………………………………………………

……………………………………………………………………………………………………………………………………

……………………………………………………………………………….

---------------




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WEEK VII

MICROBIOLOGY OF GENITOURINARY SYSTEM

Topic : Laboratory examination of organisms associated with urinary tract and sexual

transmitted infections

I. General objective

After finishing this activity, the student will be able to:

Understood microbiological examinations to confirm the diagnosis of urinary tract and

sexual transmitted infections

II. Specific objective

At the end of laboratory practice, the student could:

1. Understood methods of specimen collection and the laboratory examinations to

confirm the diagnosis of urinary tract and sexual transmitted infections

2.

Understood the methods of examination, isolation, and identification of bacteriathat cause infection of the urinary tract and sexual transmitted infections

Upon completion of this course, the student will be able to:

1. List examples of urinary and genital tract specimens.

2. Explain methods of urine sampling and genital specimen collection for microbiology

examination

3. Explain the use of the primary plating media for each specimen.

4. Describe urine culture technique for the diagnosis of UTI

5. Select incubation atmosphere, temperature, and time for each culture.

6.

Interpret, and evaluate direct Gram stains of genital specimens.

7. Describe colonial morphology and growth characteristics of the bacteria

8. Determine appropriate biochemical tests or adjunct procedures required for

identification of significant isolates.

9. Interpretation antibiotic susceptibility tests as required and evaluate their

appropriateness with regard to the treatment

10. Explain microscopic examination for diagnose Treponema pallidum infection

11. Explain the procedure and principle laboratory diagnosis of Chlamydia trachomatis

infection : microscopic, direct antigen detection, PCR

12. Correlate culture results with clinical history and presentation.




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III. Methods

Presentation

Demonstration

Discussion

Laboratory Activity

The laboratory examination is an integral part of the Microbiology course. It provides the

student an opportunity to learn basic microbiological techniques, and introduces him/her to

a case based approach to microorganisms related to respiratory tract infection.

Steps in microbiology examinations:

• Direct – detect infectious agent

–

Visualization – Culture

– Nucleic acid (DNA or RNA)

– Antigen detection

• Indirect – detect host’s response to infection

– Serological – detect antibody

Direct detection:

o Visualization (Microscopic) - smears from lesions

o Light microscope and Gram stain, wet mount preparation

Bacteria Fungal spores

o Electron microscope

Viruses

o Quick and easy if positive , negative results not definitive

Visualize bacteria:

– secretion : genital discharge

– mucosal smears : mucosal scraping

Direct detection:• Culture and identification

– Growth medium : colony count, selective media

– Adequate incubations conditions

• Temperature

• Atmosphere (O2, CO2)

– Biochemical tests for identification

• Expensive and time-consuming, positive results may be definitive

• Inoculate enrichment or selective media, incubate 24 hours

• Streak on agar plates, incubate 24 hrs

• Pick presumptive colonies and restreak for identification




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METHODS OF URINE COLLECTION

1. Random collection taken at any time of day with no precautions regarding

contamination. The sample may be dilute, isotonic, or hypertonic and may contain

white cells, bacteria, and squamous epithelium as contaminants. In females, the

specimen may contain vaginal contaminants such as trichomonas, yeast, and during

menses, red cells.

2. Early morning collection of the sample before ingestion of any fluid. This is usually

hypertonic and reflects the ability of the kidney to concentrate urine during

dehydration which occurs overnight. If all fluid ingestion has been avoided since 6

p.m. the previous day, the specific gravity usually exceeds 1.022 in healthy

individuals.

3. Clean-catch, midstream urine specimen collected after cleansing the external

urethral meatus.

4. Catherization of the bladder through the urethra for urine collection is carried out

only in special circumstances, i.e., in a comatose or confused patient. This procedure

risks introducing infection and traumatizing the urethra and bladder, thus producingiatrogenic infection or hematuria.

5. Suprapubic transabdominal needle aspiration of the bladder. When done under ideal

conditions, this provides the purest sampling of bladder urine. This is a good method

for infants and small children.

Method of clean-catch, midstream urine specimen collection:

1. Local disinfection of the meatus and adjacent mucosa should be performed with a

nonfoaming antiseptic solution; this region should then be dried with a sterile swab

to avoid mixture of the antiseptic with urine.

2.

Contact of the urinary stream with the mucosa should be minimized by spreadingthe labia in females and by pulling back the foreskin in uncircumcised males.

3. A midstream urine is one in which the first half of the bladder urine is discarded and

the collection vessel is introduced into the urinary stream to catch the last half. The

first voided specimen should be discarded since the initial urine flushes urethral

contaminants such as contaminating cells and microbes from the outer urethra. It is

the second, midstream sample that should be sent to the laboratory. This sounds

easy, but it isn't (try it yourself before criticizing the patient).

Urine collection of baby




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URINE CULTURE

1. QUALITATIVE URINE CULTURE

As previously discussed, the most common cause of UTI in men and women are gram

negative rods from the patient’s fecal flora; therefore, a qualitative approach in

demonstrating their presence should center around growth characteristics and biochemistry

associated with that group. The Qualitative isolation of urinary pathogens begins with

primary culture on Blood agar. Most all incriminating microorganisms will grow voraciously

on sheep blood with E. coli representing 80% of UTI, and many producing beta hemolytic

zones. Mac Conkey’ agar is utilized to further selectively isolate and differentiate those gram

negative rods that ferment lactose (e.g. E.coli , Klebsiella, Enterobacter and Serratia).Patients who have high numbers of cells/ ml that grows on blood agar but not on

MacConkey ‘s agar may suggest a more serious invasive pathogen of the descending route

(e.g. S. pyogenes , Candida or S. aureus).

CULTURE PROCEDURE

Step 1. Label the bottom of a blood agar and MacConkey agar plate “Qualitative

Urine”

Step 2. From the simulated urine, streak for isolation onto the blood and

MacConkey’s agar.

Step 3. Incubate plates inverted at 370C for 24 hours. Step 4. Record observations in results




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2. QUANTITATIVE URINE CULTURE

a. Pour plate method : isolation of bacteria by dilution techniques

b. Calibratied loop inoculation

A quantitative culture can be easily performed using a 1:1000 ml (0.001 ml) sterile,

disposable loop. The loop is dipped vertically into the mixed urine specimen and inoculated

onto a blood agar plate making a single line streak from the top of the plate to the bottom

of the plate. Distribution of the 1:1000 ml urine dilution is done by cross-streaking the initial

line of inoculation over the surface of the plate to obtain countable colonies. The plate is

incubated overnight at 350 C and examined the next day for countable colonies. The number

of colonies present is multiplied by 1000 (1000 represents the reciprocal of your dilution

factor “RDF” i.e. from the standardized loop 1/1000). Multiplying the number counted “NC”

by the (RDF) gives the number of organisms/ml of urine.

CFU/ml = NC x RDF

CULTURE PROCEDURE

Step. 1. Label the bottom of a blood agar plate “Quantitative Urine”

Step 2. Each desk is provided with 1/1000 ml standardized inoculating loops.

Aseptically remove one loop from the container

Step 3. Using the standardized loop, recover 1/1000 ml of urine from the

simulate specimen provided.

Step 4. Perform a quantitative culture streak

Step 5. Incubate plate at 370C for 24 hr.

Step 6. Record CFU/ml

c. Dip slide method

The functional unit consists of a clear plastic dual-chambered "paddle" containing both a

general purpose and a differential medium, Trypticase soy agar (TSA) and Levine eosin

methylene blue (EMB) agar or other kind of media, respectively. It is housed in a clear

plastic screw-cap vial. The media beds are parallel and face the same direction; each has a

surface size of about 310 mm2. The outer wall of the media wells is constructed to preventmedium dislodgement during handling and shipment and the trapping of urine between the

wall and media. To inoculate the unit, the media portion is momentarily dipped into the

urine to wet the entire agar surfaces. It is then returned to the vial and incubated in an

upright position overnight at 370C. The number of colonies per milliliter of urine is estimated

by comparing the density of growth with a standard chart obtained.




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Microorganisms causing urinary tract infections

The bacteria that cause UTI will vary according to the clinical and epidemiological setting are

usually originating from the patient’s own normal intestinal organisms; colonic flora

(coliforms = Enterobacteriaceae) which, in certain circumstances, are able to pass from the

area around the anus into the urethra and so into the bladder. Members of this bacterial

group include opportunistic pathogens (e.g. E.coli by far the most frequent cause of urinarytract infection), Klebsiella, Enterobacter, Serratia, and Proteus. Pseudomonas and

Enterococcus are also often incriminated, especially in hospitalized patients with indwelling

urinary catheters.

Community acquired UTI, otherwise healthy adults

Escherichia coli most common

Staphylococcus saprophyticus (usually young adult females)

Less commonly:

Proteus species Enterobacter species

Serratia species

Morganella species

Klebsiella species

Citrobacter species

Pseudomonas aeruginosa

Enterococcus species

(Patients with recurrent UTIs, urinary tract abnormalities, obstruction and neurogenic

bladder will have a higher frequency of those bacteria listed as ‘less common’ above).




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Hospital acquired UTI

Hospitalized patients rapidly become colonized by the hospital’s resident microflora,

specifically Gram negative bacteria. Patients in this setting are prone to UTI because they

are debilitated and may have had urinary tract instrumentation, particularly urinary tract

catheterization. Indwelling urinary catheters universally become colonized by bacteria.

Hospital bacteria have been selected by broad spectrum antibiotics used in the hospital, and

are generally quite resistant to antibiotics.

Causes of hospital acquired UTIs include:

E. coli and Enterobacteriaceae as described above.

Increased frequency of Proteus, Enterobacter , Serratia, Morganella.

Pseudomonas aeruginosa and other Pseudomonas species.

Enterococcus faecalis and E . faecium.

Stenotrophomonas maltophilia

Corynebacterium urealyticum

Staphylococcus epidermidis

Candida albicans and other species.

Uncommon causes of UTI

Some other microorganisms which may infect the urinary tract should be mentioned:

Mycobacterium tuberculosis – renal TB; uncommon in this community.

Adenovirus type 11 – “haemorrhagic cystitis” in children.

SEXUALLY TRANSMITTED INFECTIONS

N. gonorrhoeae

Specimen collection : Dacron or rayon swab

Specimen transport

1. Direct inoculation ~ Candle jars

2. Nonnutritive transport media ~Amies medium (12 hrs) should not be refrigerated

3. Nutritive transport systems ~ Transgrow, JEMBEC, Bio-Bag, Gono-Pak

Transgrow Medium is a convenient flask containing MTM Chocolate agar and CO2.

Culture

Selective Media for N. gonorrhoeae are Thayer-Martin (Modified=MTM), New York

City Medium (NYCM)

Incubation condition 35oC, 3-5% CO2

Colonial morphology : 5 types autolysin

N. gonorrhoeae forms small, convex, smooth, grayish-white to colorless, mucoid colonies in

48 hours at 35-370C.

The gonococcus requires: enriched medium with increased CO2 tension for growth

Cultured on modified Thayer Martin (MTM) Chocolate agar.

http://www.cat.cc.md.us/courses/bio141/labmanua/lab16/chocng.html














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MTM Chocolate agar is selective for pathogenic Neisseria contains :

enrichment factors to promote the growth of gonococci,

antibiotics to inhibit normal body flora :

vancomycin to inhibit gram-positive bacteria;

colistin to inhibit gram-negative bacteria;

trimethoprim to suppress Proteus;

nystatin to inhibit yeast.

Identification tests

1. Oxidase test

Oxidase

tetramethyl-ρ-phenylenediamine dihydrochloride indophenol (blue)

2. Carbohydrate utilization tests

Cystine trypticase agar (CTA) ~ 1% carbohydrate, phenol red

Organism Glucose Maltose Lactose Sucrose

N. gonorrhoeae

N. meningitides

N. lactamica

+

+

+

-

+

+

-

-

+

-

-

-

Chlamydia trachomatis

Chlamydia testing is available by 3 methods:

Culture : cell culture

Antigen detection from genital specimen: DFA. Elisa

Molecular technique: Molecular Probe, PCR

Chlamydia Culture Collection

1. Utilizing a sterile swab (rayon or dacron), obtain a suitable specimen.

2. Place the swab in transport media (M4 media – tube with pink liquid).

3. Label the transport media vial.

4. Transport the vial to the laboratory. May be refrigerated.

Chlamydia DFA/ Elisa Collection

The Direct Fluorescent Antibody (DFA) method for Chlamydia utilizes an antibody to detect

Chlamydia organisms on a slide, while Elisa detect chlamydia antigen using microtiter plate.

This method can be used for urethral, cervical, and rectal samples.

Urethral samples (male):

Patient should not have urinated one hour prior to sampling.

1. Insert small dacron swab (mini-tip) 2-4 cm into urethra.

2. Rotate swab and withdraw.




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Cervical samples (swab):

1. Wipe exocervix with large or small rayon or dacron swab to remove all excess mucus.

Dispose of swab.

2. Gently insert rayon or dacron swabs into endocervical canal until most of dacron tip is not

visible.

3. Rotate swab 5-10 seconds inside endocervical canal.

4. Withdraw swab without touching any vaginal surfaces.

Rectal samples:

Samples should be collected only from symptomatic patients.

1. Insert the large dacron swab about 3 cm into anal canal.

2. Move swab from side to side to sample crypts.

3. Withdraw swab. If fecal contamination occurs, discard swab and obtain another

specimen.

Treponema pallidum

Laboratory Diagnosis: This relies heavily on clinical manifestations. In addition, the finding of

Treponema pallidum within exudative lesions by the use of dark-field microscopy and

positive serology aids the diagnosis.

Penicillin treatment eradicates all stages, including congenital infection in pregnancy.

PROCEDURE

Step 1. Observe Prepared Slide Treponema pallidum. Special stain or negative stain

(bright field microscope) Step 2. Observe Dark Field Microscopy (presumptive Identification)

Step 3. Record results

TERMS AND QUESTIONS FOR STUDY

UTI exercise

1. What is bacteriuria?

2. How do microorganisms enter the urinary tract?

3. What group of microorganisms appear in ascending UTI’s.

Neisseria gonorrhea exercise1. Gram stain

2. Bacteria isolation, media

3. Colony morphology

4. Identification procedure

Spirochaeta exercise

1. Special stain

2. Differentiate morphology of:

Treponema pallidum

Borrelia bergdorferi Leptospira interrogans




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ADDITIONAL QUESTION

1. What is the significance of small pin point, beta hemolytic, and colonies appearing

on blood agar but no growth on MacConkey agar when conducting a qualitative

urine culture?

2.

What are significant numbers for a UTI when conducting a quantitative urine

culture? How many CFU/ml would you conclude the number of colonies if you find

145 colonies on the plate when you inoculate using a 1/1000 ml calibrated loop?

What number of colonies if you use a 1/100 ml calibrated loop?

3. Why is urine specimens keep cold until plated?

4. What do we learn from a dip stick test? Is this test conclusive?

5. Can we distinguish between N. gonorrhoeae and N. meningitidis by Gram stain?

6. What is the meaning of intracellular Gram negative cocci? Explain!

7. Why are selective media needed in primary isolation from female urogenital

specimens?

8.

Can we diagnose syphilis from Gram stained smears of primary lesions? Explain!9. Explain dark field microscopy and its application in the Microbiology laboratory!

DRAW THE RESULT OF YOUR ACTIVITY




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Signature of trainer




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WEEK VIII

PHARMACOLOGY OF GENITOURINARY SYSTEM:

DRUGS ACTING ON THE K IDNEY ; C OMPARISON

BETWEEN SOME DIURETICS AGENTS

LEARNING OBJECTIVES

After performing laboratory activity, the students should be able to:

1. Explain pharmacological properties of diuretics related to the clinical applications

RESOURCE PERSON

1. Kuswinarti (Pharmacology)

2. Ike Husen (Pharmacology)

REFERENCES

1. Benowitz. N.L. Antihypertensive agents. Katzung B.G (editor). In: Basic and

Clinical Pharmacology. 10 th Edition. McGraw Hill LANGE. 2007: 159-179

2 Ives.H.E. Diuretic agents. Katzung B.G (editor). In: Basic and Clinical

Pharmacology. 10 th Edition. McGraw Hill LANGE. 2007: 236-252

INTRODUCTION

Diuretics are drugs that increase the excretion of Na+ and water from the body by an action

on the kidney. Their primary effect is to decrease the reabsorption of Na+ and Cl- from the

filtrate, increase water loss being secondary to the increased excretion of NaCl. This can be

achieved by a direct action on the cells of the nephron or indirectly modifying the content of

the filtrate. Since a very large proportion of the NaCl and water that passes into the tubule

in the glomerulus is reabsorbed, a small decrease in reabsorption can result in a marked

increase in excretion.

Before activity, the students have to accomplish the homework assignment first, read the

primary references, so that they will be more ready in performing laboratory activity.

MATERIALS, EQUIPMENTS AND EXPERIMENTAL SUBJECTS

MATERIALS:




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a. Diuretic Agents

a. HCT

b. Furosemide

c. Spironolacton

b. Other :

d. Glycerin

e. Aquadest

EQUIPMENTS :

f. Beaker glass

g. Measuring glass

h. Catheter

i. Fixation board

j. Disposable injection

k.

NGT

EXPERIMENTAL SUBJECTS :

Three months old healthy male rabbits with the same body weight

The rabbits are divided into 2 group :

a. Control group will receive aquadest only

b. Treatment group will receive diuretic agents consist of HCT/ Furosemid

/Spironolacton.

PROCEDURES

a. Fix the rabbit on the fixation board

b. Put the catheter into the ostium urethra externum

c. Emptying the vesica urinaria

d. Administer orally 15 cc of water to the first rabbit as the control and 15 cc of

diuretic agent to the other (each diuretic type for different rabbit).

e.

Collect the urine after 30 minutes and repeat it twice every 30 minutes.f. Compare the volume of urine from :

i. Control group vs treatment group

ii. Treatment group vs treatment group




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RESULT :

Group Drug

Volume (ml)

before

treatment

Volume (ml) after treatment

30 minutes 60 minutes 90 minutes 120 minutes

I

II

III

IV

V



DISCUSSION :

CONCLUSION :

HOMEWORK ASSIGNMENT

A. DIURETICS

1. Which is the most potent diuretic found in your experiment? Give the rational

explanation of your experiment! (regarding either its mechanism of action or

pharmacokinetic)

2. List the indications of furosemide! Explain the rational explanation regarding its

mechanism of action!

3. Explain the side effects of furosemid

4. Explain the indication/s of HCT and its related mechanism of action!

5. Explain the mechanism of action of spironolacton

B. ANTI HYPERTENSIVE AGENTS

Explain the best drug of choice and worst choice for each patient below!

1. A-50-year old well controlled type 2 diabetes, male patient recently has been

diagnosed as stage II essential hypertension with normal renal function (no

microalbuminemia).

2. Hypertensive patient with tachycardia and chronic open angle glaucoma

3. Hypertensive patient with history of vasospatic angina pectoris

4.

Stage II essential hypertension patient with pregnancy.5. Patient who tends to have exacerbated asthma. Mention antihypertensive agent

should be avoided and explain the mechanism!

Lab Manual GUS

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