Case Study
Pathogenic Bacteriology
2009
Case 26
Tsung-Han Hsieh
Omar Ahmed
Rochelle Songco
Case Summary
• Patient: 80-year-old female had a cystocele
repair performed. She was give a 7-day course
of oral cephalexin but stopped the course after
3days.
• Symptoms: 10 days postoperatively, she had
diarrhea (multiple, watery, loose stool w/o blood)
for 3days. She also had crampy abdominal pain,
vomiting, and mild fever.
Case Summary
• Blood cell count was normal, but 53% of immature
polymorphonuclear cells were seen.
• Urine culture revealed >100,000 CFU/ml of E. coli which
was susceptible to all antimicrobial agents
• Cultures for Salmonella, Shigella, Yerinia, and
Campylobacter spp were all negative.
• Enzyme immunoassay (EIA) showed positive for
bacterial toxin in the stool sample.
• Physical examination, electrolytes, liver enzymes, and
lipase were within normal range.
Key Information Pointing to
Diagnosis
• >100,000 CFU/ml of E. coli in the urine
culture -> CFU/ml greater than 100,000 is
significant
• Watery diarrhea without blood -> the
organism is not invasive
• EIA showed positive for bacterial toxin in
the stool sample -> the organism produces
enterotoxin
The Diagnosis for Case #26
• The causative organism was enterotoxigenic E. coli
(ETEC)
• Cystocele repair is predisposing factor. Since E. coli is a
major cause of urinary tract infection, and the cystocele
repair involves urinary tract, the organism could have the
chance to get into the patient and cause diarrhea and
other symptoms.
• EIA was performed to specify which types of E. coli was
causing the disease.
Classification,Gram Stain Results, and
Microscopic Appearance of ETEC
• Family: Enterobacteriaceae
– Genus: Escherichia
• Gram-negative bacilli
• Facultative anaerobes
Picture was taken from http://www.nirgal.net/graphics/e_coli.jpg
Diseases and Pathogenesis of
Disease Caused by ETEC
• Disease
– E. coli is a major cause of urinary tract infection,
gastroenteritis, neonatal meningitis, and
septicemia.
» Urinary tract infections can lead to acute cystitis (bladder
infection) and pyelonephritis (kidney infection)
– ETEC is a common cause of traveler’s diarrhea
and diarrhea in children in developing countries.
» Symptoms normally include watery diarrhea, nausea,
abdominal cramps ,and low-grade fever for 1-5 days.
Diseases and Pathogenesis of
Disease Caused by ETEC
• Pathogenesis of Disease
• Pili, capsule, outer membrane proteins, and O, H, and
K antigens
• Enterotoxins – it’s produced by ETEC and causes
watery diarrhea.
–
LT (heat-labile) toxin- very similar to cholera toxin in both structure and mode of action.
It binds binds to specific Gm1 gangliosides on the epithelial cells of the small intestine
where it ADP-ribosylates Gs which stimulates adenylate cyclase to increase production
of cAMP. Increased cAMP then causes ion imbalance that results in fluid transport into
the bowel (watery diarrhea)
–
The ST (heat-stable) enterotoxin- a family of toxins which are peptides of molecular
weight about 2,000 daltons. Their small size explains why they are not inactivated by
heat. ST causes an increase in cyclic GMP in host cell cytoplasm. This also leads to
secretion of fluid and electrolytes resulting in watery diarrhea.
Diseases and Pathogenesis of
Disease Caused by ETEC
• Pathogenesis of Disease
– Particularly problematic as a nosocomial
pathogen
• It’s because E. coli is one of the M.O. that is
abundant in the hospital, and it can be transmitted
easily by fecal oral route.
Diagnosis/Isolation/Identification/ of
ETEC
•Diagnosis
•The diagnosis is suggested by the clinical picture and confirmed
by stool/urine culture. Serotyping and tests for virulence factors
are occasionally performed for outbreaks.
•Isolation
•MacConkey agar (pink to red color with or without a zone of
precipitated bile)
•ChromAgar ECC plates (light to dark blue)
•Identification
•Direct probe testing
•EIA or ELISA
Therapy, Prevention and Prognosis
of Patient Infected with ETEC
• Therapy
– Most infected persons will recover within a few days,
without requiring any specific treatment
– Electrolyte fluids are recommended for persons with
diarrhea to prevent dehydration and loss of electrolytes.
– Antibiotic course- only be give when the symptoms
become severe. ETEC is usually susceptible to a variety of
chemotherapeutic agents, but is frequently resistant to
common antibiotics such as trimethoprimsulfamethoxazole and ampicillin.
• It is essential to do susceptibility testing.
Therapy, Prevention and Prognosis
of Patient Infected with ETEC
• Prevention
– Wash your hands thoroughly after going to the
bathroom or changing diapers.
– Eat only thoroughly cooked ground beef.
– Cook ground beef products to an internal temperature
of 160 degrees Fahrenheit.
– Avoid unpasteurized milk and juices.
– Wash fresh fruits and vegetables thoroughly before
eating.
– Keep raw meat separate from ready-to-eat foods.
Therapy, Prevention and Prognosis
of Patient Infected with ETEC
• Prognosis
– The disease is usually self-limiting. The infection will
end on its own and is rarely life-threatening.
– Treatment failures might due to incomplete antibiotic
course.
– Disease relapses frequently because E. coli is NF in
the GI tract and it might turn into pathogen when the
condition is right.
Primary Research Article Contributing to the
Understanding of the Disease caused by ETEC
• Seung Y. Koh, et al., Porcine intestinal epithelial cell lines as a new
in vitro model for studying adherence and pathogenesis of
enterotoxigenic Escherichia coli, Veterinary Microbiology, 2008. 130 :
191–197.
• Experimental set up
– Bacterial adherence assays
• E. coli (different strains) were cultured overnight on 5% sheep blood agar at 32 ℃ .
• Bacteria suspension, cell lines (IPEC-J2, IPEC-1, and INT-407), and D-Mannose were
added into the plate wells.
• Phase contrast microscopy and bright field microscopy under oil were used for
observing the binding interaction.
– Green fluorescence protein-tagged E. coli
• E. coli was transformed with Green fluorescence protein (GFP).
• Bacterial adherence assays was performed with GFP tagged E. coli.
• Observation was made by confocal scanning laser microscopy
Primary Research Article Contributing to the
Understanding of the Disease caused by ETEC
•
What did they find
–
–
•
Representative figure
(A) Shows the degree of E. coli binding to
porcine intestinal epithelial cells.
(B) The confocal image showed GFPtransformed bacterial (ETEC strain B41)
attachment to IPEC-J2 cells.
(C) ETEC F18 strain 2134 intensively bound
to IPEC-1cells.
Conclusion
 Different strains of E. coli has different
binding affinity to different cell lines.
Fig. 1. Adherence of E. coli strains to IPEC-J2 and IPEC-1 cells
 Stronger binding affinity could result in releasing of more enterotoxin.
 Porcine intestinal epithelial cell lines can be
used as in vitro study of pathogenesis of ETEC.
How does this article relate to and support the case?
–
This article relates and supports the case by discussing the relationship between the binding of
ETEC to the host cell and the releasing of enterotoxin.
Take Home Message
• Disease involves
– Traveler’s diarrhea and diarrhea in children in developing
countries
• Typical symptoms are
– Water diarrhea, nausea, abdominal cramps ,and low-grade fever
• Pathogen is
– Enterotoxin: LS and ST
• Diagnostics include test
– Stool/urine culture, EIA and Serotyping for virulence factor
• Therapy is based on
– Premixed oral rehydration solutions, sometimes antibiotic course
Take Home Message
• Prognosis
– The disease is usually self-limiting and rarely life
threatening
• Prevention is
– Keep hands clean, don’t eat raw meat, and wash raw
vegetable thoroughly before eating
• Transmission is
– Via fecal-oral route or direct contact of contaminants.
• Threat is
– Major endemic health threat in developing countries
References
• Connie R. Mahon, Donald C. Lehman, George Manuselis.
Diagnostic Microbiology; 3rd ed.;Saunders, an imprint of Elsevier
Inc.:2006; Chapter 22.
• Seung Y. Koh, Sajan George, Volker Bro¨zel, et al, Porcine
intestinal epithelial cell lines as a new in vitro model for studying
adherence and pathogenesis of enterotoxigenic Escherichia coli.
Veterinary Microbiology, 2008. 130 : 191–197.
• Robin McKenziea, Michael Darsleyc, Nicola Thomasc, et al. A
double-blind, placebo-controlled trial to evaluate the efficacy of PTL003, an attenuated enterotoxigenic E. coli (ETEC) vaccine train, in
protecting against challenge with virulent ETEC. Vaccine, 2008, 26:
4731–4739
• McQueen, Nancy. Winter 2009. Lec 5 Enterobacteriaceae
Acknowledgments
• Dr. Nancy McQueen
• Omar Ahmed
• Rochelle Songco