2006 Update

Laboratory Guideline for

Ordering Stool Test for Investigation of
Suspected Infectious Diarrhea
This guideline has been developed by an Alberta Clinical RECOMMENDATIONS
Practice Guidelines Program working group and is based
on current scientific evidence. The opinions of laboratory
physicians, family physicians, pediatricians, internists, Stool testing may be required for patients with
gastroenterologists, public health and infectious disease diarrhea. The following recommendations should
physicians were used in the preparation of this guideline. guide the ordering of stool bacterial cultures (C & S),
The recommendations in this guideline reflect geographic stool ova & parasite (O & P) tests and C. difficile
and seasonal considerations in Alberta related to
toxin tests:
infectious diarrhea. This guideline was last reviewed in
2002.
♦ A CLINICAL HISTORY should be provided to
the laboratory, including the type and duration of
GOALS
symptoms, underlying medical conditions, recent
travel and recent or current antibiotic therapy.
These guidelines are intended to assist practitioners
with the following:
♦ A SINGLE stool test should be initially ordered
where indicated (see algorithm.)
♦ To improve the accuracy of stool testing for
infectious diarrhea.
♦ Stool C & S and/or stool O & P tests are usually
♦ To improve convenience for patients and
not clinically indicated for patients with onset of
parents.
diarrhea - 4 days after hospitalization (see
♦ To implement the practice of a single initial
algorithm.)
stool test sample.
♦ Consultation with an appropriate specialist is
EXCLUSIONS recommended in circumstances where additional
stool tests may be useful. Additional stool test(s)
These guidelines may not apply to the following: may be done if there is continued suspicion of
enteric bacterial infection when an initial sample
♦ Patients involved in a community or hospital is negative, and recent travel to areas where
outbreak. E. histolytica, Giardia lamblia or helminth
♦ Food handlers to whom Public Health infections are common.
regulations apply.
♦ Where an infectious etiology is not a
consideration.

The above recommendations are systematically developed statements to assis practitioner and patient decisions about testing
that may be important for clinical management of specific clinical circumstances. They should be used as an adjunct
to sound clinical decision making.
BACKGROUND
DEFINITIONS ENTERIC BACTERIAL INFECTIONS
1. Stool C & S Patients with bacterial enterocolitis typically present
with the acute onset of diarrhea and may have
Stool culture for the isolation and identification of associated fever, crampy lower abdominal pain and
enteric bacterial pathogens requires collection of an tenesmus. Although bloody diarrhea is more common
adequate stool sample, planting of the specimen when enterocolitis is caused by enteroinvasive
onto a number of selective and differential media, pathogens such as Shigella spp., not all such patients
incubation of the plates one or more times, selection with these infections will develop haematochezia. A
of appropriate colonies from incubated plates for stool C & S test must be done to diagnose suspected
identification, and in a limited number of cases, the enteric bacterial infection definitively.
provision of an antimicrobial susceptibility result.
Acute bacterial enterocolitis is acquired by ingestion
2. Stool O & P of the organism in food(s) and/or water contaminated
by feces of an infected animal or person. Commonly
Stool examination for the diagnosis of enteric parasitic implicated food sources include raw and undercooked
colonization and infection involves submitting a stool eggs and egg products, raw milk and milk products,
specimen in fixative to the laboratory, preparation meat, salads and poultry and their products. Direct
of the stool for staining and examination (i.e., contact with an infected animal (e.g., pet turtles or
concentration and/or filtration), and staining with a chicks) may also transmit Salmonellosis. The incidence
standard stain such as iron haematoxylin. of enteric bacterial infections is highest in travellers
to underdeveloped countries where food handling
3. C. difficile Toxin practices and sanitary conditions may be poor.
Epidemics may also occur in families, in groups
Stool testing for diagnosis of antibiotic associated attending a restaurant or social function where a
colitis involves submitting a stool specimen in a common contaminated food source is ingested, or
sterile container to the laboratory, for detection of in institutions such as daycare centres and nursing
C. difficile toxin(s) using a variety of different assay homes.
methods. There is controversy about the optimal
method of toxin(s) detection. Currently, the three In developed areas of the world, a limited number of
major assay methods include enzyme immunoassay bacterial pathogens are responsible for acute bacterial
for toxins A and B, latex agglutination assays for enterocolitis, as outlined in Table 1.
toxins A and B, and cell culture cytotoxicity assays
with specific neutralization of toxin B. Culture of
C. difficile from stool is not considered diagnostic Table 1. Enteric Bacterial Pathogens
since a significant number of children and adults carry
this organism as part of their normal colonic flora. • Campylobacter jejuni
• Salmonella spp.
4. Enteric Viral Infections • Verotoxin producing E. coli
(e.g., E. coli O157:H7)
Enteric viral infections are diagnosed by submitting • Shigella spp.
a stool sample in a sterile container using a variety • Aeromonas spp. (toxin producing strains)
of different methods depending upon the type of • Clostridium difficile
virus(es) being sought. Although rapid antigen • Yersinia enterocolitica
detection testing for Rotavirus infection in children is • Pleisiomonas shigelloides
widely available, most other viral tests are only done
by the Provincial Public Health Laboratories in
Calgary and Edmonton.
It remains controversial whether all or only toxin
producing strains of Aeromonas spp. from stool
samples are pathogenic.
Vibrio spp. infections are uncommon but are usually symptoms alone cannot distinguish between enteric
reported in travellers to underdeveloped countries infection with protozoa versus bacteria. Some patients
where sanitary conditions are poor (e.g., Vibrio who have recently travelled to or immigrated from
cholerae), or in people eating contaminated shellfish under-developed countries may have mixed infections.
(e.g., Vibrio parahaemolyticus). Microscopic examination of stool remains the main
technique for confirming the presence of enteric
In Alberta, enteric infections occur year-round but parasitic infection, although antigen detection
have a definite peak in the summer months. This techniques may assume a greater role in the future.
likely happens because of improper food handling
practices which are associated with outdoor activities. Colonization and/or infection with protozoa is
Infections which occur at other times of the year are therefore most common in people who have travelled
more likely to have been acquired while travelling in to an underdeveloped country where sanitary
underdeveloped countries. conditions may be poor, in children attending day care
centres, in institutionalized patients, and in homosexual
Table 1 reflects the order of incidence of pathogens. men.
Campylobactor infections are the most common
enteric disease in Alberta. There is also no difference Although all enteric protozoa encountered in a given
in the pattern or frequency of isolation of the types stool specimen may be reported, not all may require
of bacterial pathogens isolated from patients admitted specific antimicrobial treatment (Table 2).
to the hospital with a primary diagnosis of acute
enterocolitis, versus those attending out-patient areas
including the emergency room.1,2,3 Table 2. Pathogenic Enteric Protozoa

C. difficile is the main cause of antibiotic associated • Giardia lamblia (intestinalis)

diarrhea and pseudomembranous colitis. C. difficile • Cryptosporidium spp.
has become one of the most commonly detected • Entamoeba histolytica
enteric pathogens, particularly in hospitalized and • Dientamoeba fragilis
nursing home patients where infection may be • Microsporidium spp.
nosocomially transmitted. Symptoms range from • Blastocystis hominis
mild diarrhea to the most severe form of the disease, • Cyclospora spp.
toxic megacolon, where bowel perforation may • Isospora belli
require total colectomy. Rarely, death may occur if
appropriate treatment is not given. Although microbial
agents of all classes may cause this problem, the most
commonly implicated agents include ampicillin, Although the role of Blastocystis hominis in causing
clindamycin and cephalosporins. C. difficile toxin symptoms is controversial, it may be pathogenic in
mediated colitis may also rarely occur in patients who patients where no other cause is found for diarrhea.
have not recently received antibiotic treatment. Other enteric protozoa not listed above may frequently
colonize the bowel, but have not been documented to
Physicians should take a clinical history before cause diarrhea or other gastrointestinal symptoms even
ordering stool cultures. History should include in immunocompromised patients.
questions about recent travel, other risk factors for
acquiring bacterial infections and exposure to other Although intestinal helminth infections rarely cause
people with similar symptoms. diarrhea, stool O & P tests are also used to diagnose
these infections including roundworms (nematodes)
ENTERIC PARASITE INFECTION such as hookworm, Strongyloidiasis, Ascariasis etc.,
tapeworms (cestodes) such as Taenia spp.,
Patients with enteric infection with protozoan Hymenolepsis spp., etc. and flukes (trematodes)
parasites may present with persistent diarrhea such as Schistoso-miasis, Clonorchiasis etc. Because
(i.e., ≥ 5 days duration). Intestinal parasite infection the exact number of stool examinations needed to
may also cause abdominal bloating and flatulence, but diagnose enteric helminth infections is not known,
additional stool samples for examination (i.e., 2 or ♦ Underlying disease(s) - immunosuppression
3 sequential samples) may be required. Three ♦ Recent travel history (Dates/Location)
investigations may also be required if E. histolytica, ♦ Recent or current antibiotic therapy
or Giardia lamblia infection is suspected.
1. Enteric Bacterial Infections
Enteric Viral Infections
Microbiology laboratories routinely inoculate stool
Viral gastroenteritis occurs predominately in two C & S specimens onto a variety of selective and
distinct clinical forms. The first pattern of disease is differential medias and into broth which enhances the
typified by Rotavirus which primarily causes severe, isolation of all major enteric bacterial pathogens of
acute watery diarrhea in infants and young children interest. Special planting media to recover Vibrio spp.
each year during the winter months, although sporadic will only be inoculated when symptomatic patients
cases can occur at other times during the year. have recently travelled to an endemic area for V.
Rotavirus infection may also be transmitted amongst cholerae infection, or have recently ingested shellfish
hospitalized children, and nosocomial Rotavirus (i.e., suspected V. parahaemolyticus infection). A
outbreaks are well described. Enteric adenovirus history of recent or current antibiotic use should be
40/41 infections are a much less common cause provided when ordering C. difficile toxin testing.
of pediatric diarrhea but sporadic cases may be Most microbiology laboratories only perform toxin(s)
reported. Caliciviruses and astroviruses may also testing and do not do stool cultures for the organism.
cause diarrhea in children, but in general, these viruses
are associated with milder diarrheal symptoms. 2. Enteric Parasitic Infections
Calicivirus infection may be more prevalent in children
in day care. Enterovirus infections most often present Routine stool staining and examination will detect
as aseptic meningitis and rarely cause diarrheal illness. most enteric parasites of interest except for
The second pattern of illness is typified by Norwalk Cryptosporidium spp., Microsporidium spp. and
virus and is characterized by evolving community-wide Cyclospora spp. Special stool concentration and
outbreaks which affect one or more family members staining methods are required to detect these
as well as their contacts. Most symptomatic viral infections. The microbiology laboratory may only
gastroenteritis illness in adults occurs within this perform these tests if an appropriate history is
setting, and outbreaks in institutionalized elderly provided (i.e., child in a day care centre, HIV
patients have been documented. Table 3 outlines seropositive patient, recent travel to the tropics or
the most common types of enteric viral infections farm exposure). In addition, recent or current
diagnosed in Alberta in order of decreasing frequency antibiotic therapy may decrease the laboratory’s
from left to right. ability to detect enteric parasite infection.

3. Enteric Viral Infections

Table 3. Enteric Viral Infections
Laboratory testing for enteric viral infections is not
• Rotavirus • Astroviruses usually necessary except in young children with acute,
• Adenovirus 40/41 • Norwalk virus severe acute diarrheal illness during the late fall and
• Caliciviruses • Norwalk-like viruses winter months when Rotavirus infection becomes
epidemic each year. Rapid antigen detection tests such
as latex agglutination or enzyme immunoassay are
CLINICAL HISTORY widely available for Rotavirus infections. Although
Adenovirus 40/41 infection occurs much less
A clinical history is essential for the efficient and frequently, it can also be diagnosed using an enzyme
appropriate work-up of stool C & S and stool O & P immunoassay. Electron microscopy and stool viral
samples. The following information must be provided cultures must be performed to diagnose other types
on the requisition: of enteric viral infections.
♦ Clinical symptoms and duration
♦ Type of suspected infection/exposure
RESEARCH FINDINGS 4 children with Dientamoeba fragilis, 2 children with
Blastocystis hominis and 2 children with
Basis of the Multiple Sample Rule Cryptosporidiosis.

Until recently, 3 sequential stool samples (i.e., different However, due to the retrospective design of the study,
bowel movements) for stool C & S and stool O & P the clinical significance of any of these cases in terms
testing have usually been ordered for diagnosis of of patient symptoms or institution of treatment could
enteric bacterial and/or parasitic infection respectively. not be ascertained.
Historically, the Centre for Disease Control (CDC)
in the United States recommended the practice Evidence for Not Performing Stool C & S or
of performing multiple sequential stool O & P Stool O & P Tests on Hospitalized Patients
examinations based upon studies which showed (i.e., ≥ 4 days in hospital)
that the rate of recovery of E. histolytica from
asymptomatic patients increased from 50%, with It has been well documented that there is a very low
only a single stool sample examination to 90%, after yield of finding enteric infections (i.e., bacterial or
examination of 6 sequential stool samples.4 Several protozoal) with onset ≥ 4 days after hospitalization
other studies have also reported that multiple stool regardless of the age of the patient or their immune
specimens are required to achieve adequate sensitivity status.1,7-11 Stool C & S and stool O & P tests
of recovery of parasites on examination of stool are therefore not recommended in hospitalized patients
specimens.5,6 However, these studies were except in rare situations where the patient has recently
epidemiological in nature, aimed at primarily travelled to an underdeveloped area, or where an
diagnosing asymptomatic E. histolytica excretion in additional stool examination is required for follow up
patients in the underdeveloped world, and stool of a previously diagnosed infection.
concentration methods were not always used.
It has become clear that these guidelines may not Most diarrhea in hospitalized patients is due to
be applicable or appropriate for patients in the C. difficile toxin-mediated colitis when antibiotics
developed world where there is not a high incidence have been recently given, i.e., symptoms may occur up
of E. histolytica infection. There has also not been to 8 weeks after antimicrobial(s) were stopped. Viral
scientific evidence to support the routine ordering gastroenteritis, particularly Rotavirus infection, is a
of multiple sequential stool C & S samples, and common cause of diarrhea in children during the late
this practice was likely extrapolated from the fall and winter months and infection may be
recommendations for stool O & P testing. nosocomially transmitted in hospitalized children.
Other causes of viral gastroenteritis such as
Evidence for the Single Sample Rule Adenovirus 40/41 or other enteric viruses are much
less common but should be considered when other
Recent, predominantly adult studies have documented types of infection have been ruled out.
the high sensitivity of a single stool culture for enteric
bacterial pathogens and stool examination for parasites Experience with Implementation
in a large general hospital setting.7-11 Recently,
reported work from the Alberta Children’s Hospital Since these recommendations were implemented at
has also demonstrated the high efficiency for diagnosis the Alberta Children’s Hospital more than 2 years
of enteric infections of a single initial stool culture and ago, there has been a sustained reduction in both stool
stool parasite examination in both hospitalized and C & S and stool O & P testing. Physicians have been
ambulatory children.1,11 Most pediatric cases of very supportive of the initial rule of a single sample,
enterocolitis (190 of 194, 98%) are confirmed from and these types of stool tests are now rarely
a single stool culture, and a second sample is seldom requested for patients with prolonged hospital stays.
required.1,3 Most clinically relevant protozoal Nursing personnel and parents also appreciate not
infections (102 of 112, 91%) were also detected in the having to collect multiple stool samples.
first stool specimen examined.3 Infections which
would have been missed on a single stool O & P
specimen included 4 children with Giardiasis,
ADVICE TO PATIENTS 10. Asnis DS, Bresciani A, Ryan M, McArdle P, Mollura JL,
Ilardi CF. Cost-effective approach to evaluation of
diarrheal illness in hospitals. J CLIN MICROBIOL
Clear communication by physicians and other health 1993;31:1675.
care personnel to the patient and/or parents is integral 11. Kabani A, Cadrain G, Trevenen CL, Jadavji TJ, Church
to their understanding and compliance with the need DL. Practice guidelines for ordering stool ova and
parasite testing in a pediatric population. AM J CLIN
to test for diarrhea pathogens. It is important to PATHOL 1995;104:272-278.
explain the nature of their infection, the expected
duration of diarrhea and to outline the management Toward Optimized Practice (TOP)
plan. The proper stool sample collection methods Program
for different types of tests (i.e., stool C & S, stool
O & P, C. difficile, Rotavirus) should be outlined to
Arising out of the 2003 Master Agreement, TOP succeeds the former
ensure that the samples will be of acceptable quality Alberta Clinical Practice Guidelines program, and maintains and
to the laboratory (see patient collection instruction distributes Alberta CPGs. TOP is a health quality improvement
sheet). initiative that fits within the broader health system focus on quality
and complements other strategies such as Primary Care Initiative and
the Physician Office System Program.
An important part of managing patients with infectious
diarrhea is to tell them how they can remain well The TOP program supports physician practices, and the teams they
hydrated throughout their illness. For patients with work with, by fostering the use of evidence-based best practices and
confirmed bacterial enterocolitis, the reasons for not quality initiatives in medical care in Alberta. The program offers a
variety of tools and out-reach services to help physicians and their
routinely treating the illness with antibiotics should be colleagues meet the challenge of keeping practices current in an
explained. The potential side effects of antimicrobial environment of continually emerging evidence.
therapy must be outlined to patients (parents of
children) with severe bacterial enterocolitis or
pathogenic enteric protozoal infections who receive TO PROVIDE FEEDBACK
antibiotics. Patients should also be informed that
Public Health staff may contact them about what they The Alberta CPG Working Group for Microbiology is
have been eating or drinking to try to identify the a multi-disciplinary team composed of microbiologists,
source of a notifiable infection. general practitioners, and a gastroenterologists,
pathologist, university representative, member of the
SELECTED REFERENCES public and representative of Alberta Health and Wellness.
The team encourages your feedback. If you need more
1. Church DL, Cadrain G, Kabani A, Jadavji T, Trevenen C. information or if you have difficulty applying this guideline,
Practice guidelines for ordering stool cultures in a please contact:
pediatric population. AM J CLIN PATHOL
1995;103:149-153
2. Siegel DL, Edelstein PH, Nachamkin I. Inapproprite Toward Optimized Practice Program
testing for diarrheal diseases in the hospital. JAMA 12230 - 106 Avenue NW
1990; 263:979-982. EDMONTON, AB T5N 3Z1
3. Fan K, Morris AJ, Reller LB. Application of rejection T 780. 482.0319
criteria for stool cultures for bacterial pathogens. TF 1-866.505.3302
J CLIN MICROBIOL 1993;31:2233-2235. F 780.482.5445
4. Alicna AD, Fadell AJ. Advantage of purgation in E-mail: [email protected]
recovery of intestinal parasites or their eggs. AM J Infectious Diarrhea - March 1997
CLIN PATHOL 1959;31:139-142.
5. Andrews J. The diagnosis of intestinal protozoa from Reviewed 2006
purged and normally passed stools. J PARASITOL
1934;20:253-254.
6. Morris AJ, Wilson ML, Reller LB. Application
of rejection criteria for stool ovum and parasite
examinations. J CLIN MICROBIOL 1992;30:3213-3216.
7. Koplan J, Benfari Ferraro MJ, Fineberg HV, Rosenberg
ML. Value of stool cultures. LANCET 1980;23:413-416.
8. Montessori GA, Bishcoff L. Searching for parasites in
stool: once is unually enough. CMAJ 1987;137:702.
9. Senay H, MacPherson D. Parasitology: diagnositc yield
of stool examinations. CMAJ 1989;140:1327-31.
 Algorithm for Investigation of Suspected Infectious Diarrhea

History & Physical Exam

Duration < 5 days Duration ≥ 5 days

Child under 14 and: All patients: All patients:

• Severe Watery Diarrhea and/or Toxic; or Loose/Watery Diarrhea Severe, Bloody Diarrhea
• Hospitalized for > 4 days; or (Mild to Moderate) and and/or Patient Toxic(b)
• Daycare Not-Toxic(a)

Recent Antibiotic Use(c) Recent Antibiotic Use(c) Recent Antibiotic Use(c) Recent Antibiotic Use(c)

Yes No Yes No No Yes No Yes

C. difficle Viral C. difficle Wait Stool C. difficle Stool C & S x 1 - C. difficle

toxin test
- Tests(d) toxin test No Test C&Sx1 toxin test (if not done toxin test
- -
& Stool before) & Stool
- > - C&Sx1 O & P x 1(e)

If Diarrhea Specialized Tests

persists ≥ 5 days, defined by
+ + + follow Algorithm + + Consultation + +

Appropriate Clinical Management(f)

Notes:
a) Some cases of watery diarrhea in these patients do not require investigation and are self limiting.
b) Bacterial enterocolitis is most often acquired in Alberta from late Spring (May/June) to Fall (October/September). Cases outside of these peak months occur most frequently in travellers/
immigrants from developing countries.
c) Antibiotic associated colitis (AAC) due to C. difficile may occur up to 8 weeks after antibiotics have been stopped. AAC is the most common cause of diarrhea in hospitalized patients.
d) Rotavirus infections occur predominantly in young infants/children (≤ age 3 years) and are epidemic in Alberta each year from early Winter (November/December) to early Spring (April/May).
Rotavirus infections may also be nosocomially transmitted between hospitalized children. Adenovirus 40/41 and other enteric viral infections occur much less frequently throughout the year.
e) Multiple, sequential stool ova and parasite examinations may be necessary for patients who have recently travelled and/or immigrated (i.e., usually with the past 6 months) from an underde-
veloped country in order to diagnose E. histolytica, G. lamblia and/or enteric helminth infections.
f) Rehydration is central to the management of patients with infectious diarrhea.
March 1997
Laboratory Appendix for
Ordering Stool Tests for Investigation of
Suspected Infectious Diarrhea

This appendix complements the Guideline for Toward Optimized Practice

Ordering Stools Tests for Investigation of
Suspected Infectious Diarrhea prepared by the (TOP) Program
Microbiology Working Group of the Clinical
Practice Guidelines Program. Arising out of the 2003 Master Agreement, TOP
succeeds the former Alberta Clinical Practice Guidelines
RECOMMENDATIONS program, and maintains and distributes Alberta CPGs.
TOP is a health quality improvement initiative that fits
within the broader health system focus on quality and
♦ Use containers with enteric solution for stool
complements other strategies such as Primary Care
culture and sensitvity specimens which Initiative and the Physician Office System Program.
require transportation of greater than two
hours from the site of collection to the The TOP program supports physician practices, and
laboratory. the teams they work with, by fostering the use of
evidence-based best practices and quality initiatives in
♦ Use containers with SAF fixative for parasite medical care in Alberta. The program offers a variety of
testing. tools and out-reach services to help physicians and their
colleagues meet the challenge of keeping practices
current in an environment of continually emerging
♦ Use normal sterile containers for:
evidence.

• stool culture and sensitivity tests where

transport of the specimen for a period of
greater than two hours from collection Toward Optimized Practice Program
site to laboratory is not required; 12230 - 106 Avenue NW
EDMONTON, AB T5N 3Z1
T 780. 482.0319
• testing for C. difficile; or TF 1-866.505.3302
F 780.482.5445
• testing for Rotavirus and other enteric E-mail: [email protected]
viruses.

♦ Relevant clinical history should be provided

to properly process the specimen.

♦ In patients with AIDS, laboratories may

choose to look routinely for Microsporidiosis,
Cryptospridiosis and Mycobacterium avium
complex (MAC).

♦ Many of the infectious agents mentioned

in the Guideline, “Ordering Stool Tests for
Investigation of Suspected Infectious
Diarrhea,” are classified as notible to Public
Health.