Lab Diagnosis of GI and Intra-abdominal Infections

Laboratory Diagnosis of GI and
Intra-abdominal Infections
Dr. Brad Cutrell
October 30, 2014

GI and Intra-abdominal
Infections
• IDSA Guidelines and recent
NEJM review article on
Infectious Diarrhea
• IDSA Diarrhea Guidelines
(Clin Infect Dis 2001 ; 32:331 -350.)
• IDSA Intra-abd Infxn Guidelines
(Clin Infect Dis 2010 ; 501 : 133 -164.)
• Focus of this talk on often
overlooked topic of diagnostic
testing and stewardship in
case based format
DuPont HL. NEJM 2014; 370:1532-40.

Tenets of Specimen Management
• Tenets with particular relevance to this discussion:
1. Inappropriate or poor quality specimens should be rejected.
- Formed stool sent for diarrheal evaluation
2. Physicians should not demand a report of “everything that
grows.”
- GI sites heavily populated with normal GI flora.
3. Lab requires a specimen, not a swab of a specimen.
- Adequate sample of fluid or tissue vital for culture recovery.
4. Specimens should be labeled accurately and completely to
allow reliable result interpretation.
- Precise site and method of collection important to
distinguish colonization from clinically relevant bacteria.

Case #1: Esophagitis
• 35 yo WM with PMHx of AIDS
(last CD4 25, not on HAART)
presents with 2 week history of
burning substernal chest pain and
pain on swallowing. Despite 4
days of fluconazole, symptoms
persist and patient undergoes
EGD which reveals the following
(see image).
• What is the DDx?
• What are the most appropriate
lab Dx tests?

AIDS-related Esophageal Disease
• Reflux esophagitis or drug-related common in immunologically normal host
• Infectious causes higher in immunosuppressed patients
• Often empiric trial of anti-fungals (esp. if thrush present) with EGD if not
improvement within 72 hours
Disease Entity Characteristic Features
Fungal
Candida esophagitis “Cottage cheese”; oral thrush; entire esophagus
Ulcerative
CMV esophagitis
HSV esophagitis
Idiopathic or aphthous ulcers
Deep, “punched out” ulcers; distal 1/3
Widespread, shallow erosions; Oral/labial lesions
Similar to CMV
Neoplastic
KS or lymphoma
Esophageal carcinoma
Constitutional symptoms; Esophageal mass lesion

Reflux esophagitis
CMV esophagitis
HSV esophagitis
Candida esophagitis

Esophagitis: Dx Testing
• Diagnosis is made by endoscopic appearance and histopathologic
evaluation of biopsy for typical features
• Fungal culture for Candida spp. if resistant species suspected
• Viral culture for HSV primarily, less useful for CMV
IDSA Guide to Utilization of Micro Lab. Clin Infect Dis 2013; 57(4):e22-121.

Case #2: Gastritis
• 54 yo male with PMHx of recently diagnosed peptic ulcer
disease presents to clinic. He was seen at OSH for bleeding
gastric ulcer 3 months ago and treated for 8 weeks with high
dose PPI, which is now stopped. He is asymptomatic but
wonders if additional testing is needed. No biopsy records are
available but he was not given any abx.
• What diagnostic testing is available to evaluate for
Helicobacter pylori?

Helicobacter pylori
• Indications for testing
- Active or Hx of PUD - Functional dyspepsia (not GERD)
- MALToma or Gastric Cancer - ITP, ? Unexplained B12 or IDA
Dx Test Sens/Spec Affected by
PPI/Abx/GIB?
Test of Cure?
Invasive
Rapid urease test
Histology
Culture
90%/95%
90%/98%
73%/100%
X
+/-
X
Not usually since
requires EGD
Noninvasive
Urea breath test
Stool Ag
Serology
> 90%/95%
95%/> 90%
85%/80%
X
X
X
X
• Delay Dx Testing: D/c Acid Rx- 2 wks, D/c Abx- 4 wks, GIB- 4-8 wks
• Test of Cure: UBT or Stool Ag > 4 wks after Rx
• Low PPV for serology in areas with low incidence and only detects exposure

Case #3: Diarrhea Part 1
• 34 yo WF with no PMHx presents with 5 days of diarrhea and
abdominal pain. She reports 6 BMs per day. She denies fevers
but reports some blood in her stools as well as mucus. She
works as a daycare employee. She ate at a church picnic 10
days ago, but is not aware of anyone else from there with
diarrhea. She also took augmentin for 5 days for a “sinus
infection” 4 weeks ago.
• What is your DDx?
• What diagnostic evaluation is appropriate?
• What empiric abx would you give her while you wait?

Diarrhea Testing Algorithm
IDSA Infectious Diarrhea Guidelines. Clin Infect Dis 2001; 32: 331-50.

Routine Stool Testing
• What plates are set up for a routine stool Cx?
• Blood Agar plate, MacConkey Agar plate, Campy
agar, XLD (selective differential media)
• What routine enteric pathogens are detected?
• Shigella spp., Salmonella spp., Campylobacter spp.
• What additional tests would be indicated here?
• ELISA assay for Shiga-toxin production
• Sorbitol MacConkey plate for E. coli O157:H7
• C. difficile testing

Routine Stool Testing
• More than 1 stool culture is rarely indicated (87-94% of
pathogens detected on first sample in adults)
• NAAT testing being developed but most not yet FDA-approved
or commercially available
• What if the patient from case had eaten shellfish?
• Special culture for Vibrio spp.
• What if she had iron overload from chronic transfusions?
• Special culture for Yersinia spp.
• Other pathogens requiring specialized stool testing:
• Aeromonas, Plesiomonas, Edwardsiella, Staph aureus, various E.
coli (ETEC, EIEC, EPEC, EAEC)
• Toxin detection for outbreaks related to Bacillus cereus,
Clostridium perfringens and C. botulinum

• 55 yo AAF admitted to hospital with acute pyelonephritis, with
blood and urine Cx positive for E. coli sensitive to FQ. She is
clinically improving on levofloxacin, but on day 4 of her
hospital stay, she develops recurrence of fevers, abdominal
pain and new onset non-bloody, watery diarrhea (5 episodes
in 24 hrs).
• What is the appropriate diagnostic workup?

Nosocomial Onset Diarrhea
• “3 Day Rule”
• In patients who develop hospital-onset diarrhea (≥3 d), yield for stool
Cx and O&P is very low; thus, testing is NOT generally recommended
• Represents 15-50% of specimens submitted ($20-75 mil USD 1996)
• HIV or other immunocompromised state (neutropenia) exception
• Conversely, 15-20% of specimens from patients with hospital-onset
diarrhea (≥ 3 d) are positive for C. difficile
• Don’t forget about non-infectious causes of diarrhea in
hospitalized patients (medications, tube feedings, etc.)
IDSA Infectious Diarrhea Guidelines. Clin Infect Dis 2001; 32: 331-50.

Dx Testing: C. difficile
Goal: Detect presence of toxigenic strains of C. difficile actively
producing toxin leading to infection
Test Sens Spec Time to
Result
Comments
ELISA for GDH
Ag
High Low Rapid Does not distinguish toxigenic and
non-toxigenic strains
ELISA for toxin
A/B
Low High Rapid Only 70-85% sensitive
PCR for toxin
genes
High High Rapid Expensive; Cannot distinguish
active infection from carriage
Toxigenic
culture
High High Very slow Gold std but slow and labor
intense
Cytotoxin
assay
High High Slow Similar limitations to Cx
Current preferred testing strategies include either 1) PCR or 2)
two-step testing with ELISA for GDH Ag and Toxin
Solomon D, et al. Open Forum ID (Spring 2014); E-pub 4/2/2014.

3 D’s of C. difficile Testing
• Don’t send formed stool for testing!
• Don’t repeat a negative PCR test for at least 7 days!
• Don’t check a repeat “test of cure”!
• When in doubt, consider calling in Cliff:
Coming to a ward near you soon !?
Cliff the C. diff Dog

• 32 yo HM with PMHx of advanced AIDS (last CD4 25, not on
HAART) presents to the hospital with low grade fevers and
profuse watery diarrhea. The patient reports diarrhea for the
past 2 weeks, 8-10 watery BMs daily. He has diffuse
abdominal pain but no peritoneal signs. No gross blood or
mucus in stools. He has lost 15 lbs. and feels generally weak.
Labs show K 2.8, Mg 1.2, Cr 1.4. Routine stool Cx is negative.
• What is the DDx for the cause of diarrhea?
• What diagnostic workup is appropriate?

Parasitic Stool Evaluation
• Evaluation for persistent diarrhea, esp. in IC
hosts, warrant expanded testing including for
parasites
• Traditional direct microscopy required timely
transport (< 4h) of fresh stool and often
multiple samples to ↑ yield
• Modern kits with preservatives increase time
window but may affect ability to do special
stains or immunoassays
• Most cost-effective to only send additional
samples if 1st negative AND patient remains
symptomatic

Dx Testing: Select Pathogens
Pathogen O&P Micro Stool EIA Select Comments
E. histolytica X X Indistinguishable from E.
dispar on micro
Blastocystis hominis X Role as pathogen unclear
Dientamoeba fragilis X PCR available
Giardia lamblia X X Giardia/Crypto combo EIA
Cryptosporidium X* X *Modified acid fast stain
Cyclospora X* *Modified acid fast stain
Cystoisospora X* *Modified acid fast stain
Microsporidia X# #Modified trichrome stain; EM
on small bowel Bx
Worms X Forms seen based on life cycle
MAC/MAI AFB blood and stool Cx, Bx

• 52 yo WF s/p DDKT 8 months prior is admitted with 6 days of
fevers, abdominal pain and diarrhea. Diarrhea is watery, 6-8
BMs daily and occult blood positive. CT scan shows bowel
wall thickening in the colon diffusely. Patient is D+/R-,
received ATG induction Rx and is on maintenance tacrolimus,
prednisone, and mycophenolate. Routine stool Cx and C.
difficile testing is negative.
• What is the DDx?
• What diagnostic testing is appropriate for the most likely
etiology?

CMV GI Disease
• CMV tissue invasive GI disease usually
seen in patients with advanced AIDS, IBD,
or SOT/HSCT patients
• Serum or plasma CMV PCR not sensitive or
specific enough to confirm or exclude
diagnosis of CMV colitis
• Lower endoscopy preferred diagnostic
modality
• Perform multiple biopsies for
histopathology, IHC stains and viral Cx (if
resistance a concern)

Other GI Viruses
• Rotavirus
• Rapid detection EIA Ag testing commercially available
• Calicivirus (Norovirus)
• NAAT (PCR) testing available via public health or reference labs
• Enteric Adenovirus
• EIA Ag testing available via reference labs
• Viral stool Cx and NAAT testing available for other viruses, but
strongly encourage discussion with Micro attending if desired

Case #7: Proctitis
• 23 yo MSM presents with rectal pain and bleeding as well as
tenesmus and rectal discharge. He had an outside flex
sigmoidoscopy and was told he likely has early IBD based on
random biopsies showing granulomatous proctitis. He is
sexually active with multiple partners, has history of multiple
STIs in past. Previously lived in the Netherlands, but has
moved to US for work.
• What diagnoses should be considered?
• What diagnostic workup should be performed?

Proctitis DDx
de Vries, et al. Int J STD AIDS. 2014 Jun; 25 (7): 465-74.

Proctitis Diagnostic Workup
Pathogen Diagnostic Testing
N. gonorrhoeae Rectal swab or bx NAAT*, Gram stain and Cx
C. trachomatis Rectal swab or bx NAAT*; Serology/genotyping for LGV
HSV Rectal swab or bx NAAT or viral culture
Syphilis Serum RPR with confirmatory test; Dark field
microscopy or multiplex NAAT from ulcer (not widely
available)
Enteric pathogens
and parasites
Stool Cx and stool O&P; Histopathology
CMV Typical histopathology with IHC stains on biopsy
* Though widely used, rectal swab is not FDA-approved specimen for GC NAAT.
de Vries, et al. Int J STD AIDS. 2014 Jun; 25 (7): 465-74.

Case #8: Liver Abscess
• 60 yo Taiwanese male with poorly
controlled DM2 presents with 4
weeks of fevers and dull RUQ pain.
He lives in US but spends his
summers hiking and traveling
through South America and
volunteering as a sheep-herder. His
CT scan of his abdomen is shown.
• What is the DDx and what diagnostic
workup is appropriate?

Liver Abscess
Etiology Diagnostic Testing
Pyogenic Liver Abscess
(E. coli, Klebsiella, Strep
anginosus group, Staph
aureus, anaerobes,
polymicrobial)
Blood Cx x 2
Gram stain and aerobic/anaerobic culture from
aspirated material
Amebic liver abcess
(E. histolytica)
Imaging (US or contrast CT scan)
Serology or serum Ag (high NPV if endemic)
Liver aspirate antigen and direct microscopy
Hydatid cyst disease
(Echinococcosis)
Imaging (US)
Serology
Direct visualization of aspirated material

Diagnostic Testing for Amebiasis
Stool O&P and antigen testing useful for diarrhea, but
NOT hepatic disease
Haque R, et al. N Engl J Med 2003; 348:1565-73.

Case #9: Peritonitis
• 52 yo Hispanic female originally from Mexico with PMHx of
cryptogenic cirrhosis presents with increasing ascites and
abdominal pain. She is afebrile and otherwise appears well
without complaints. A diagnostic paracentesis is performed
which shows WBC 850 with 75% PMNs.
• What diagnostic workup should be performed?

Peritonitis Workup
Etiology Microbiology Dx Testing
Spontaneous bacterial
Monomicrobial, usually
peritonitis
aerobic enterics
• 10-50 cc peritoneal fluid for
centrifugation and aerobic Cx
• Consider direct inoculation of
aerobic blood Cx bottle
• Blood Cx, ? UA and urine Cx
Secondary peritonitis Polymicrobial, including
anaerobes and Candida
• 10-50 cc fluid for aerobic and
anaerobic Cx
• Strongly consider abd imaging
• Blood Cx
What about direct inoculation of peritoneal fluid into blood Cx?
• One study showed significant improvement from 76% to 100% yield with
immediate blood Cx inoculation vs delayed Cx
• Sterile technique using new needle to inoculate Cx is recommended
• Do NOT directly inoculate blood Cx if secondary peritonitis is suspected
Runyon BA, et al. J Clin Microbiol 1990; 28(12):2811.

Peritonitis: Special Scenarios
• What if the same case presentation but now ascites fluid
showed WBC of 550 cells with 85% lymphs?
• Tuberculous peritonitis: Notoriously difficult Dx to make
- AFB smear and Cx (< 20%); Consider large volume tap (250-500 cc) with
centrifugation to increase yield (Coordinate with micro lab!)
- Fluid MTB PCR and adenosine deaminase (best in non-cirrhotics)
- Laparoscopic peritoneal biopsy
- Evaluation for pulmonary or disseminated disease
• Fungal peritonitis: fluid fungal stain and Cx, systemic workup
• Malignant ascites: fluid cytology, imaging with laparascopic or
image guided biopsy
• PD-associated peritonitis: evaluation is similar but microbiology
is very different: GPC > GNR, must consider indolent pathogens
such as fungal, mycobacterial including NTMs

Conclusions: Dx Stewardship
• Don’t send formed stool to lab for diarrheal evaluation
• Don’t reflexively send multiple stool samples for Cx or O&P unless
patient remains symptomatic
• Don’t routinely send stool Cx or O&P for hospital onset diarrhea (≥ 3
days)
• 3 D’s for C. difficile (Don’t send formed stool, Don’t repeat within 7
days, Don’t check test of cure)
• Don’t ask the lab to workup every organism from non-sterile GI sites
• Pursue endoscopy and tissue diagnosis if CMV is a concern

Lab Diagnosis of GI and Intra-abdominal Infections

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