Case Study #1: E. coliCase Study #1: E. coli

Tania GuevaraTania Guevara

AnthonyAnthony

EdmundEdmund

March 11, 2009March 11, 2009

California State University of Los AngelesCalifornia State University of Los Angeles

Case SummaryCase Summary Age Age -19 yr old female-19 yr old female

Medical HistoryMedical History -History of urinary tract infection (UTI)-4 mts prior to -History of urinary tract infection (UTI)-4 mts prior to

admissionadmission -Treated with oral ampicillin-Resistance to ampicillin can -Treated with oral ampicillin-Resistance to ampicillin can

occur because the oral medication is rapidly excreted and occur because the oral medication is rapidly excreted and

the duration of significant drug concentration in the urine the duration of significant drug concentration in the urine is short. An additional reason is that ß-lactams are is short. An additional reason is that ß-lactams are relatively ineffective in clearing gram negative rods from relatively ineffective in clearing gram negative rods from the vaginal and colonic mucosa, thus possibly the vaginal and colonic mucosa, thus possibly predisposing to recurrences when used to treat UTIpredisposing to recurrences when used to treat UTI

Case SummaryCase Summary SymptomsSymptoms Five Days Prior to AdmissionFive Days Prior to Admission -Nausea w/out vomiting-Nausea w/out vomiting One Day Prior to AdmissionOne Day Prior to Admission -Left flank pain-Left flank pain -Fevers (38.8oC)-Fevers (38.8oC) -Chills-Chills -Increased urinary frequency-Increased urinary frequency -Foul smelling urine-Foul smelling urine

Physical examinationPhysical examination - Left costovertebral angle tenderness- Left costovertebral angle tenderness UrinalysisUrinalysis -Notable for >50 WBC/ high power field, 3-10RBC/ high power -Notable for >50 WBC/ high power field, 3-10RBC/ high power

field, and 3+ bacteriafield, and 3+ bacteria -Urinary culture was positive for >100,000 CFU of an org/ml-Urinary culture was positive for >100,000 CFU of an org/ml

Case SummaryCase Summary

>50 WBC>50 WBC -Pyuria refers to the presence of abnormal numbers of -Pyuria refers to the presence of abnormal numbers of

leukocytes that may appear with infection in either the leukocytes that may appear with infection in either the upper or lower urinary tract. White cells from the vagina, upper or lower urinary tract. White cells from the vagina, especially in the presence of vaginal and cervical infections, especially in the presence of vaginal and cervical infections, may contaminate the urine. If two or more leukocytes per may contaminate the urine. If two or more leukocytes per each high power field appear in non-contaminated urine, each high power field appear in non-contaminated urine, the specimen is probably abnormal. the specimen is probably abnormal.

3-10 RBC3-10 RBC -Theoretically, no red cells should be found, but some find -Theoretically, no red cells should be found, but some find

their way into the urine even in very healthy individuals. their way into the urine even in very healthy individuals. However, if one or more red cells can be found in every However, if one or more red cells can be found in every high power field, and if contamination can be ruled out, the high power field, and if contamination can be ruled out, the specimen is probably abnormal. specimen is probably abnormal.

Case SummaryCase Summary 3+ bacteria3+ bacteria

-Bacteria are common in urine specimens because of the -Bacteria are common in urine specimens because of the abundant normal microbial flora of the vagina and because abundant normal microbial flora of the vagina and because of their ability to rapidly multiply in urine standing at room of their ability to rapidly multiply in urine standing at room temperature. temperature.

-Diagnosis of bacteriuria in a case of suspected urinary tract -Diagnosis of bacteriuria in a case of suspected urinary tract infection requires culture. A colony count may also be done infection requires culture. A colony count may also be done to see if significant numbers of bacteria are present. to see if significant numbers of bacteria are present. Generally, more than 100,000/ml of one organism reflects Generally, more than 100,000/ml of one organism reflects significant bacteriuria. significant bacteriuria.

Key Information Pointing to Key Information Pointing to DiagnosisDiagnosis

UTI are a bacterial infection of the urinary bladder (cystitis) UTI are a bacterial infection of the urinary bladder (cystitis) or of the kidneys (pyelonephritis)or of the kidneys (pyelonephritis)

Nearly all UTI’s are caused by bacteria and enter the Nearly all UTI’s are caused by bacteria and enter the urethral opening and move upward to the urinary bladder urethral opening and move upward to the urinary bladder and sometimes the kidneysand sometimes the kidneys

Possible pathogens for a UTI:Possible pathogens for a UTI: -E.coli-E.coli -enterobacteriae Klebsiella-enterobacteriae Klebsiella -P. mirabilis-P. mirabilis -P. aeroginosa-P. aeroginosa -Enterococci (group D)-Enterococci (group D) -Staphylococcus-Staphylococcus

Key Information Pointing to Key Information Pointing to DiagnosisDiagnosis

Cystitis (lower UTI)- sudden onset, frequency, urgency, Cystitis (lower UTI)- sudden onset, frequency, urgency, burning, or painful voiding of small vol. of urine. Nocturia burning, or painful voiding of small vol. of urine. Nocturia with suprapubic and lower back pain is common. Urine is with suprapubic and lower back pain is common. Urine is turbid, gross hematuria, and pneumaturia can result. Low turbid, gross hematuria, and pneumaturia can result. Low grade fever.grade fever.

Pyelonephritis (Upper UTI)-Pyelonephritis (Upper UTI)-ChillsChills, , feversfevers, , flank painflank pain, , nauseanausea, , and vomiting. and vomiting. Costovertebral angleCostovertebral angle percussion tenderness percussion tenderness is generally present in the infected side.is generally present in the infected side.

Most common common organism causing this type of Most common common organism causing this type of infection if E. coli (uropathogenic E. coli-UPEC)infection if E. coli (uropathogenic E. coli-UPEC)

Key Information Pointing to Key Information Pointing to DiagnosisDiagnosis

UTI’s are more frequent in women because they have a shorter UTI’s are more frequent in women because they have a shorter urethra, which makes it easier for bacteria to move up the urinary urethra, which makes it easier for bacteria to move up the urinary tract and provide less barrier to the bacterial invasiontract and provide less barrier to the bacterial invasion

The Diagnosis for Case # The Diagnosis for Case # The diagnosis provides clear evidence to support that the The diagnosis provides clear evidence to support that the

patient is suffering from a pyelonephritis UTI caused by patient is suffering from a pyelonephritis UTI caused by Uropathogenic E. coli organism.Uropathogenic E. coli organism.

Classification,Gram Stain Results, and Classification,Gram Stain Results, and Microscopic Appearance of Pathogen XMicroscopic Appearance of Pathogen X

Classification-more than 15 genera Classification-more than 15 genera -Escherichia-Escherichia Morphology and General CharacteristicsMorphology and General Characteristics -Gram negative, non-sporing, rod shape bacteria-Gram negative, non-sporing, rod shape bacteria -NF of intestinal tract, some are enteric pathogens and -NF of intestinal tract, some are enteric pathogens and

others are urinary or respiratory pathogensothers are urinary or respiratory pathogens -Anaerobes -Anaerobes -Oxidase (-)-Oxidase (-) -Ferment glucose and may/not produce gas -Ferment glucose and may/not produce gas -Reduce nitrate to nitrite-Reduce nitrate to nitrite -If motile-by peritrichous flagella-If motile-by peritrichous flagella

Classification,Gram Stain Results, and Classification,Gram Stain Results, and Microscopic Appearance of Pathogen XMicroscopic Appearance of Pathogen X

Most grow well on a variety of lab mediaMost grow well on a variety of lab media

-Lactose fermenters:-Lactose fermenters: On CBA they all produce large and dull gray colonies. BetaOn CBA they all produce large and dull gray colonies. Beta

hemolytichemolytic Differentiation is based on biochemical reactions and Differentiation is based on biochemical reactions and

differences in antigenic structures (O, H, and K1)differences in antigenic structures (O, H, and K1)

Diseases and Pathogenesis of Diseases and Pathogenesis of Disease Caused by Pathogen XDisease Caused by Pathogen X

Virulence FactorsVirulence Factors -Toxins-Toxins• Enterotoxins (LT/ST)Enterotoxins (LT/ST)• Shiga-type toxinShiga-type toxin• Enteroaggregative ST-like toxinEnteroaggregative ST-like toxin• EndotoxinEndotoxin -Type III Secretion System-Type III Secretion System -Adhesion (pili or fimbriae and non-fimbrial)-Adhesion (pili or fimbriae and non-fimbrial) VF that protect the bacteria from hostVF that protect the bacteria from host -Capsule-Capsule -Iron capturing ability-Iron capturing ability -Outer membrane proteins-Outer membrane proteins

Virulence factors of recognized importance in the pathogenesis of urinary Virulence factors of recognized importance in the pathogenesis of urinary tract infection (UTI) include: adhesins (P fimbriae, certain other mannose-tract infection (UTI) include: adhesins (P fimbriae, certain other mannose-resistant adhesins, and type 1 fimbriae), the aerobactin system, hemolysin, resistant adhesins, and type 1 fimbriae), the aerobactin system, hemolysin, K capsule, and resistance to serum killing K capsule, and resistance to serum killing

Diseases and Pathogenesis of Diseases and Pathogenesis of Disease Caused by Pathogen XDisease Caused by Pathogen X

In women who suffer from recurrent UTI’s- suggest that this In women who suffer from recurrent UTI’s- suggest that this may be due to the formation of pod-like E. coli biofilms may be due to the formation of pod-like E. coli biofilms inside the bladder epithelial cellsinside the bladder epithelial cells

-Bacteria living on the edges of the biofilms may break off -Bacteria living on the edges of the biofilms may break off leading to a round of infection.leading to a round of infection.

Diagnosis/Isolation/IdentificationDiagnosis/Isolation/Identification

• UTI DiagnosisUTI Diagnosis

A culture of the bacteria may be done to determine the type A culture of the bacteria may be done to determine the type of bacteriaof bacteria

A A clean-catch urine specimenclean-catch urine specimen. This test involves . This test involves cleansing the area around the urethral opening and cleansing the area around the urethral opening and collecting a mid-stream urine sample, preventing bacteria in collecting a mid-stream urine sample, preventing bacteria in the genital area from contaminating the sample.the genital area from contaminating the sample.

UrinalysisUrinalysis is performed to determine the level of white blood is performed to determine the level of white blood cells that destroy harmful bacteria (leukocytes) in the urine. cells that destroy harmful bacteria (leukocytes) in the urine. A large number of these cells may indicate bacterial A large number of these cells may indicate bacterial infection.infection.

Dipstick:Dipstick: - check for appearance and color of urine- check for appearance and color of urine -pH or concentration-pH or concentration

Diagnosis/Isolation/IdentificationDiagnosis/Isolation/Identification Microscopic testMicroscopic test -Blood cells (e.g., red blood cells, white blood cells; -Blood cells (e.g., red blood cells, white blood cells; indicate indicate hematuriahematuria or infection) or infection) -Casts (e.g., hyaline, granular; may indicate kidney -Casts (e.g., hyaline, granular; may indicate kidney disease) disease) -Cells from the lining of the urinary tract (epithelial cells) -Cells from the lining of the urinary tract (epithelial cells) -Crystals (may indicate metabolic disease) -Crystals (may indicate metabolic disease) -Fat (may indicate -Fat (may indicate nephroticnephrotic syndrome syndrome or or diabetic diabetic neuropathy)neuropathy) -Renal tubular cells (may indicate acute tubular -Renal tubular cells (may indicate acute tubular necrosis)necrosis)

Therapy, Prevention and Prognosis Therapy, Prevention and Prognosis of Patient Infected with UTIof Patient Infected with UTI

TherapyTherapy -are often treated with antibacterial drugs.-are often treated with antibacterial drugs. -The type of drug used and the duration of treatment depend -The type of drug used and the duration of treatment depend

on the type of bacteriaon the type of bacteria -Most UTI’s are treated with trimethoprim-sulfamethoxazole -Most UTI’s are treated with trimethoprim-sulfamethoxazole

(e.g. Bactrim, Cotrim, Septra), amoxicillin (e.g. Amoxil, (e.g. Bactrim, Cotrim, Septra), amoxicillin (e.g. Amoxil, Trimox), or fluoroquinolones (e.g. Levaquin, Cipro). Trimox), or fluoroquinolones (e.g. Levaquin, Cipro).

-The infection may improve within a couple of days, but 1 to -The infection may improve within a couple of days, but 1 to 2 weeks of medication may be prescribed to prevent a 2 weeks of medication may be prescribed to prevent a kidney infection. kidney infection.

Note: UTIs that are caused by bacteria such as Note: UTIs that are caused by bacteria such as Chlamydia Chlamydia trachomatistrachomatis and and mycoplasma hominismycoplasma hominis require a longer require a longer course of treatment with tetracycline (e.g. Achromycin), course of treatment with tetracycline (e.g. Achromycin), trimethoprim-sulfamethoxazole, or doxycycline trimethoprim-sulfamethoxazole, or doxycycline (e.g.Periostat).(e.g.Periostat).

Prevention and Prognosis of Prevention and Prognosis of Patient Infected with UTIPatient Infected with UTI

Drink plenty of water every day.Drink plenty of water every day. Urinate when you feel the need; don't resist the urge to urinate.Urinate when you feel the need; don't resist the urge to urinate. Wipe from front to back to prevent bacteria around the anus from Wipe from front to back to prevent bacteria around the anus from

entering the vagina or urethra.entering the vagina or urethra. Take showers instead of tub baths.Take showers instead of tub baths. Cleanse the genital area before sexual intercourse.Cleanse the genital area before sexual intercourse. Avoid using feminine hygiene sprays and scented douches, which Avoid using feminine hygiene sprays and scented douches, which

may irritate the urethra.may irritate the urethra. Some doctors suggest drinking cranberry juice.Some doctors suggest drinking cranberry juice.

Prognosis:Prognosis: 12-13 cases annually per 10,000 population in women. UTIs. The 12-13 cases annually per 10,000 population in women. UTIs. The

elderly are at increased risk for such infections because the elderly are at increased risk for such infections because the bladder doesn't empty fully due to certain prostate and bladder bladder doesn't empty fully due to certain prostate and bladder conditions. conditions.

Primary Research Article Contributing to the Primary Research Article Contributing to the Understanding of the Disease caused by E. Understanding of the Disease caused by E.

colicoli Mysorekar, Indira U, et al., 2007, Mechanisms of Uropathogenic Escherichia Mysorekar, Indira U, et al., 2007, Mechanisms of Uropathogenic Escherichia

coli Persistence and Eradication From The Urinary Tract, coli Persistence and Eradication From The Urinary Tract, Proceedings of the Proceedings of the National Academy of Sciences of the United States of AmericaNational Academy of Sciences of the United States of America, 103: 14170-, 103: 14170-14175.14175.

-Study aim to demonstrate in a murine model of UTI, that UPEC established -Study aim to demonstrate in a murine model of UTI, that UPEC established quiescent intracellular reservoirs (QIR’s) in Lamp 1+ endosomes within quiescent intracellular reservoirs (QIR’s) in Lamp 1+ endosomes within urinary bladder epithelium, thus resulting in reoccuring UTI’s.urinary bladder epithelium, thus resulting in reoccuring UTI’s.

-However, treatment of infected bladder harboring QIR’s with the cationic -However, treatment of infected bladder harboring QIR’s with the cationic protein, protamine sulfate, would lead to epithelial exfoliation and protein, protamine sulfate, would lead to epithelial exfoliation and eradication of bacteria in 100% of the animals. eradication of bacteria in 100% of the animals.

Set-Up:Set-Up:o Mice- 6-8 wks female C57BL/6 mice, all maintained under pathogen free Mice- 6-8 wks female C57BL/6 mice, all maintained under pathogen free

conditionsconditionso Bacterial strains- UTI89, a UPEC strain recovered from human patient w Bacterial strains- UTI89, a UPEC strain recovered from human patient w

cystitis, transformed, and grown as a static culturecystitis, transformed, and grown as a static cultureo Inoculation of mice-Mice were anesthetized and inoculated w/ 50 ul of 10Inoculation of mice-Mice were anesthetized and inoculated w/ 50 ul of 1077

CFU bacteria (bladders removed and processed for microscopy, histology, CFU bacteria (bladders removed and processed for microscopy, histology, and CFU titration)and CFU titration)

Primary Research Article cont’dPrimary Research Article cont’d

o CFU titration-Bacterial titers were determine by plating serial CFU titration-Bacterial titers were determine by plating serial dilutions of bladder homogenatesdilutions of bladder homogenates

o BrdU labeling-some mice received an intraperitoneal injection of BrdU labeling-some mice received an intraperitoneal injection of an aq. solution of BrdU.an aq. solution of BrdU.

o Histologic and IFA analysis-bladders were dissected and cut Histologic and IFA analysis-bladders were dissected and cut longitudinally. longitudinally.

o SEMSEMo Whole mount Analysis-Mice were killed, bladders were Whole mount Analysis-Mice were killed, bladders were

bisected and sprayed with PBS to observe outline of mature bisected and sprayed with PBS to observe outline of mature facet cells and visualize nuclei.facet cells and visualize nuclei.

o Urine analysis- At indicated times, mice were induced to Urine analysis- At indicated times, mice were induced to urinate by applying gentle pressure to the skin just below urinate by applying gentle pressure to the skin just below the occiput,the occiput,

o Statistical AnalysisStatistical Analysis

Primary Research Article cont’dPrimary Research Article cont’d Findings:Findings:

1.1. QIR’s within bladder may be a source of recurrent UTI’s.QIR’s within bladder may be a source of recurrent UTI’s.

2.2. Inducing epithelial exfoliation may be a therapeutic avenue for Inducing epithelial exfoliation may be a therapeutic avenue for treating this infection.treating this infection.

Relatedness to Topic:Relatedness to Topic:

-Since pyelonephritis often succeeds initial cystitis, this particular -Since pyelonephritis often succeeds initial cystitis, this particular study can help prevent pyelonephritis from occurring by study can help prevent pyelonephritis from occurring by resolving cystitis.resolving cystitis.

Take Home MessageTake Home Message UPEC involves the organism E. coli which is a gram negative rod UPEC involves the organism E. coli which is a gram negative rod

that causes UTI.that causes UTI. Typical symptoms include chills, fevers, flank pain, nausea, and Typical symptoms include chills, fevers, flank pain, nausea, and

vomiting. Costovertebral angle percussion tenderness is generally vomiting. Costovertebral angle percussion tenderness is generally present in the infected side.present in the infected side.

Pathogen is Uropathogenic E. coliPathogen is Uropathogenic E. coli UTI diagnosis can be performed from a clean catch urine UTI diagnosis can be performed from a clean catch urine

specimen, urinalysis, and microscopic test.specimen, urinalysis, and microscopic test. Therapy is based on antibiotics that include include Therapy is based on antibiotics that include include

fluoroquinolones, amoxillin, trimethoprim, etc. fluoroquinolones, amoxillin, trimethoprim, etc. Symptoms of a bladder infection usually disappear within 24 - 48 Symptoms of a bladder infection usually disappear within 24 - 48

hours after treatment begins. If you have a kidney infection, it hours after treatment begins. If you have a kidney infection, it may take 1 week or longer for your symptoms to go away. may take 1 week or longer for your symptoms to go away.

Prevention to infection includes drinking plenty of water and Prevention to infection includes drinking plenty of water and cranberry juice, taking showers instead of baths, and good genital cranberry juice, taking showers instead of baths, and good genital hygiene.hygiene.

Nearly all UTI’s are caused by bacteria and enter the urethral Nearly all UTI’s are caused by bacteria and enter the urethral opening and move upward to the urinary bladder and sometimes opening and move upward to the urinary bladder and sometimes the kidneysthe kidneys

Pyelonephritis is very common, with 12-13 cases annually per Pyelonephritis is very common, with 12-13 cases annually per 10,000 population in women. UTIs. The elderly are at increased 10,000 population in women. UTIs. The elderly are at increased risk for such infections because the bladder doesn't empty fully risk for such infections because the bladder doesn't empty fully due to certain prostate and bladder conditions. due to certain prostate and bladder conditions.

ReferencesReferences Mysorekar, Indira U, et al., 2007, Mechanisms of Uropathogenic Mysorekar, Indira U, et al., 2007, Mechanisms of Uropathogenic

Escherichia coli Persistence and Eradication From The Urinary Escherichia coli Persistence and Eradication From The Urinary Tract, Tract, Proceedings of the National Academy of Sciences of the Proceedings of the National Academy of Sciences of the United States of AmericaUnited States of America, 103: 14170-14175., 103: 14170-14175.

Rosen, David et al, 2007, Detection of Intracellular Bacterial Rosen, David et al, 2007, Detection of Intracellular Bacterial Communities in human Urinary Tract Infection, Communities in human Urinary Tract Infection, Journal PmedJournal Pmed, , (4)12; e 329.(4)12; e 329.

Dr Mcqueen. "Enterobacteriaceae." Enterobacteriaceae. Bio 244, Dr Mcqueen. "Enterobacteriaceae." Enterobacteriaceae. Bio 244, Los Angeles. 22 Jan. 2009. Los Angeles. 22 Jan. 2009.

Mahon, Connie et al. Mahon, Connie et al. Textbook of Diagnostic MicrobiologyTextbook of Diagnostic Microbiology. 3. 3rdrd Ed. Ed. St. Louis: Missouri, 2007. St. Louis: Missouri, 2007.

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