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Accurate diagnosis of Giardia spp and proper fecal examination procedures


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A series of investigations evaluated the ability of different testing methods - a swing-head centrifugation technique using two flotation solutions (1.18-specific gravity zinc sulfate and 1.27-specific gravity Sheather's sugar solution), a passive commercial flotation technique, and the SNAP Giardia Test Kit from IDEXX Laboratories - to identify Giardia-positive dogs and recover the eggs of other intestinal parasites. It was determined that the SNAP Giardia test can improve a practice's ability to identify Giardia-infected dogs. Because of its higher specific gravity, the sugar solution was better for recovering heavy parasite eggs, such as Taenia spp, and thus is the flotation solution of choice when conducting routine centrifugation fecal examinations.
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Veterinary Therapeutics Vol. 7, No. 1, Spring 2006
*Sponsored by IDEXX Pharmaceuticals, Inc.,
Greensboro, NC.
Aseries of investigations evaluated the ability of different testing methods—a
swing-head centrifugation technique using two flotation solutions (1.18–specif-
ic gravity zinc sulfate and 1.27–specific gravity Sheather’s sugar solution), a
passive commercial flotation technique, and the SNAP Giardia Test Kit from
IDEXX Laboratories—to identify Giardia-positive dogs and recover the eggs of
other intestinal parasites. It was determined that the SNAP Giardia test can im-
prove a practice’s ability to identify Giardia-infected dogs. Because of its high-
er specific gravity, the sugar solution was better for recovering heavy parasite
eggs, such as Taenia spp, and thus is the flotation solution of choice when con-
ducting routine centrifugation fecal examinations.
Accurate Diagnosis of Giardia spp and Proper
Fecal Examination Procedures*
M. W. Dryden, DVM, PhD
P. A. Payne, DVM, PhD
V. Smith, RVT
Department of Diagnostic Medicine/Pathobiology
College of Veterinary Medicine
Kansas State University
Manhattan, KS 66506
Afecal examination is considered a routine
procedurein many veterinary practices, but in
our experience, often little thought is given to
performing the procedurecorrectly.Although
fecal flotations should be performed byveteri-
narians or trained veterinary technicians, the
assignment is commonly given to the newest
staff member, with very little instruction or
emphasis on the importance of the task pro-
vided. For the past 10 years, informal surveys
of second-year veterinary students at Kansas
State University indicate that more than 90%
of the students had conducted “fecal examina-
tions” in practices where they had worked or
volunteered, yet most of these students had no
formal training in the proper performance of a
fecal flotation or identification of parasite eggs,
oocysts, or cysts.
Accurate evaluation of fecal samples is im-
portant and must be taken seriously by all
members of the clinical practice. Not only is
pet health at stake, but several of the parasites
that may be recovered and identified are po-
tentially zoonotic, such as Toxocara canis, Tox-
ocara cati, Ancylostoma caninum,Giardia spp,
Cryptosporidium parvum,and Toxoplasma
gondii. Arecently published study demonstrat-
ed that a 5-minute swing-head centrifugation
technique could recover more parasite eggs and
M. W. Dryden, P. A. Payne, and V. Smith
and emails we receive, we realize that many vet-
erinary practices find it difficult to diagnose
this disease using fecal examinations. There are
several reasons why identifying giardiasis is dif-
ficult in private practice. Many pseudoparasites,
such as yeasts (Figure 1), plant remnants, and
debris, havebeen mistaken for these tiny or-
ganisms. Identification of Giardia cysts is fur-
ther compromised because microscopes used in
private practice areoften not equipped with
micrometers that can allowmeasurement of
cysts as small as 8 to 12 ×7 to 10 µm. Cysts are
shed intermittently,and repeated fecal analyses
may be needed beforecysts are recovered in a
sample.2,3 Cysts aredelicate and deteriorate rap-
idly in fecal flotation solutions; if a fecal exam-
ination is conducted using a solution other
than zinc sulfate (ZnSO4), the cysts may be dis-
torted (Figures 2 and 3). In many clinics, the
only technique used is the direct smear; howev-
er, trophozoites are fragile and are often found
only in very fresh, diarrheic feces. Finally, be-
cause of the difficulty in detecting Giardia spp,
veterinarians have told us that they often rely
on a patient’s response to metronidazole treat-
ment as a presumptive diagnosis.
Several studies have demonstrated that re-
covery of Giardia cysts can best be accom-
oocysts and had fewer false-negative results
than passiveflotation techniques.1While a
swing-head centrifugation technique is more
efficient than passive flotation and will signifi-
cantly aid in the diagnosis of many parasites, it
is our experience that many practices using this
technique still struggle with the accurate iden-
tification of Giardia spp.
Giardiasis is a diagnostic dilemma. Weagree
that Giardia is one of the most commonly mis-
diagnosed, underdiagnosed, and overdiagnosed
parasites. Based on the numerous phone calls
Figure 1. Yeasts (12–15 µm) recovered in a fecal ex-
amination and incorrectly identified as Giardia cysts
in a referring veterinarypractice.
Figure 2. Giardia cysts stained with Lugol’s iodine re-
covered using 1.18-SG ZnSO4in a swing-head cen-
trifugation technique.
Figure 3. Giardia cysts recovered using 1.27-SG
Sheather’s sugar solution in a swing-head centrifuga-
tion technique.
Veterinary Therapeutics Vol. 7, No. 1, Spring 2006
plished using a 1.18–specific gravity (SG)
ZnSO4centrifugation technique.2–5 This has
been the diagnostic test of choice for many vet-
erinarypractices and veterinary diagnostic lab-
oratories, including that of Kansas State Uni-
versity. While the ZnSO4centrifugation
technique recovers and maintains the integrity
of Giardia cysts more consistently than other
flotation techniques, it still does not alleviate
the problem of proper cyst identification. Sev-
eral fecal antigen tests have been developed
that appear to have high sensitivity in detecting
Giardia antigen in human feces; however, they
do not have the same level of sensitivity in de-
tecting giardiasis in dogs and cats when com-
pared with centrifugation using ZnSO4.3,4,6
This article describes several investigations
that were designed to compare the fecal antigen
SNAP Giardia T
est Kit (Idexx Laboratories) and
aZnSO4centrifugation fecal flotation tech-
nique. Additional data on centrifugation versus
apassivefecal flotation technique for the recov-
eryof parasite eggs and oocysts arealso present-
ed so that more complete recommendations can
be made concerning routine parasite diagnostic
procedures for private veterinary hospitals.
Two groups of purpose-bred beagles (14 in
2004; 12 in 2005) housed at the Animal Re-
source Facility at Kansas State University for
other research projects were found to be natu-
rally infected with Giardia spp and shedding
cysts on a routine basis. In 2004, the 14 dogs
had been allocated for the other research proj-
ect to eight runs, with two dogs in each of six
runs and one dog each in two other runs. In
2005, the 12 dogs had been allocated to seven
runs, with two dogs in each of five runs and
one dog each in two other runs. Once a week
for 4 to 5 weeks, feces was collected from the
runs and examined for the presence of Giardia
cysts. An attempt was made to collect feces
from each dog in pens that housed two dogs.
The fecal samples from a pen were mixed thor-
oughly and split into two subsamples. One
Standard Swing-Head Centrifugation Fecal Examination Technique
1. Weigh out 2–5 g of feces.
2. Mix feces with approximately 10 ml of flotation solution.
3. Pour mixture through a tea strainer into a beaker or fecal cup.
4. Pour strained solution into a 15-ml centrifuge tube.
5. Fill tube with flotation solution so that a slight positive meniscus forms.a
6. Place a coverslip on the tube, and put the tube in the centrifuge.
7. Make sure the centrifuge is balanced.
8. Centrifuge at 1,200 rpm (280 ×g)for 5minutes.
9. Remove the tube and let stand 10 minutes.
10. Remove the coverslip, and place it on a glass slide. Systematically examine the entire area under the
coverslip at 100×magnification (i.e., 10×objective). You may wish to use the 40×objective lens to
confirm your diagnosis and make measurements; however, with practice, most parasites can be iden-
tified using the 10×objective (100×magnification).
aDo not overfill the tube. Doing so will cause some of the floating eggs to be forced down the side of the tube when
the coverslip is placed.
Adapted from: Dryden MW, Payne PA, Ridley R, Smith V: Comparison of common fecal flotation techniques for the
recovery of parasite eggs and oocysts. Vet Ther 6(1):16, 2005.
subsample was evaluated with a swing-head
centrifugation technique (see box on page 6)
using 1.18-SG ZnSO4,with one drop of Lu-
gol’s iodine placed on the slide before place-
ment of the coverslip. The second subsample
was evaluated using the SNAP Giardia Test Kit
according to label directions. Samples were
scored as either positive or negative. Each fecal
sample was evaluated by one of the authors
(V.S.), who has more than 14 years of experi-
ence conducting and examining fecal samples
at Kansas State University.
Athird evaluation of the SNAP Giardia Test
Kit was conducted by second-year veterinary
students during the fall of 2005. Fecal samples
from 116 puppies were provided by a local bro-
ker. These students had just listened to a lecture
on giardiasis, been given a short visual presenta-
tion on identification of Giardia cysts, and had
previously conducted direct smears and fecal ex-
aminations using the swing-head centrifugation
technique before participating in this exercise.
Students were also given written directions on
conducting SNAP Giardia tests.
For the direct smear, a small sample of feces
was placed on a glass slide and mixed with a
drop or two of saline. The material was spread
thinly, a drop of Lugol’s iodine was added, and
the slide was covered with a glass coverslip.
Each fecal sample was analyzed using a direct
smear technique, centrifugation, and the
SNAP Giardia Test Kit. Samples and tech-
niques were recorded as either positive or neg-
ative. Each of the 107 veterinary students con-
ducted at least one fecal examination, and nine
students evaluated two samples.
Data for the final evaluation of diagnostic
techniques came from a “wet lab” conducted at
the 2005 Central Veterinary Conference. A
group of veterinarians and veterinarytechni-
cians attended a wet lab on conducting proper
fecal examinations. Participants were provided
with a fecal sample from naturally parasitized
dogs. The sample was a pooled sample from
several dogs and contained eggs of A. caninum,
Eucoleus boehmi, Taenia spp, T. canis, and
Trichuris vulpis; oocysts of Cystoisospora spp;
and Giardia spp cysts. In addition, the sample
tested positive on the SNAP Giardia fecal anti-
gen test kit.
Participants were given a short lecture before
the wet lab on how to conduct the various
techniques, provided written instructions, and
shown color images of all parasite eggs,
oocysts, and cysts that were in the fecal sample.
Visual instruction included images of Giardia
cysts recovered in ZnSO4and sugar solutions
(Figures 2 and 3). Each participant collected
two “quarter-sized” samples from the feces and
conducted a 15-minute Ovassay (Synbiotics), a
swing-head centrifuge technique (5-minute
spin at 280 ×gfollowed by a 10-minute wait
beforeanalysis; see boxon page 6), and a
SNAP Giardia test. The participants were di-
vided into two groups:
Group 1 used 1.18-SG ZnSO4for the flota-
tion solution and added one drop of Lugol’s
iodine to the slide beforeplacing the coverslip.
Modified Sheather’s Solution (SG 1.27)a
454 g granulated sugar
355 ml tap water
6ml formaldehyde
Dissolve sugar and water in the top of a dou-
ble boiler or with gentle heat. If solution is
not clear, filter it through coarse filter paper.
aCheck specific gravity (SG) with a hydrometer that
has a range compatible with the solution being test-
ed. Hydrometers with ranges of 1.000–1.400 are
Adapted from: Dryden MW, Payne PA, Ridley R,
Smith V: Comparison of common fecal flotation
techniques for the recovery of parasite eggs and
oocysts. Vet Ther 6(1):17, 2005.
M. W. Dryden, P. A. Payne, and V. Smith
Veterinary Therapeutics Vol. 7, No. 1, Spring 2006
Group 2 used 1.27-SG Sheather’s sugar so-
lution (see box on page 7).
Participants were asked to record the number
of eggs, oocysts, or cysts recovered as 0, 1 to
10, 11 to 50, or more than 50 per slide. Com-
pleted results were returned by 14 participants
in Group 1 and 13 in Group 2.
As is typical of Giardia infections, cyst shed-
ding was not consistent during the evaluation
period of the naturally infected beagles (Tables
1and 2). Fecal samples from beagles evaluated
in 2004 werefound to be positive using the
ZnSO4centrifugation method 87.5%, 75%,
37.5%, and 50% of the time during weeks 1,
2, 3, and 4, respectively (Table 1). The SNAP
Giardia fecal antigen test produced a positive
reaction in 62.5%, 50.0%, 50.0%, and 62.5%
of the fecal samples during weeks 1, 2, 3, and
4, respectively. During 2005, fecal samples
from the beagles were found to be positive us-
ing the ZnSO4centrifugation method 57.14%
of the time in week 1 and 85.57% during
weeks 2 and 3, and all were positive in week 5
(Table 2). The SNAP Giardia fecal antigen test
produced a positive reaction 85.57% of the
time in fecal samples collected during weeks 1
to 3, and all samples were antigen positive in
weeks 4 and 5 (Table 2).
Interestingly,there weretimes when one or
both tests produced false-negative results (Ta-
bles 1 and 2). In 2004, when results from both
tests werecombined, the accuracy of diagnosis
was increased during three of the four weeks.
Combining test results led to a positive diag-
nosis in 100.0%, 75.0%, 50.0%, and 75.0%
TABLE 1. Evaluation of Repeated 1.18-SG ZnSO4Centrifugation Fecal Flotations and
SNAP Giardia Tests to Identify Giardia-Positive Beagles: Trial 1, 2004
Pen No. of Week 1 Week 2 Week 3 Week 4
No. of dogs or 7 5 6 4 3 4 4 5
pens testing
Total no.of dogs 8 6 5 6
or pens testing
of the fecal samples during weeks 1, 2, 3, and
4, respectively. In the 2005 trial, however, only
during week 3 did combining test results in-
crease diagnostic accuracy (Table 3).
In the evaluations conducted by the second-
year veterinary students, almost half (56 of
116) of the puppy fecal samples were recorded
as positive for Giardia (Table 3). The direct
smear technique detected the fewest number of
positive samples, with students recording only
four positivesamples. These direct smear tech-
nique data may be artificially low since the fe-
cal samples were collected several hours before
being tested and trophozoites may have been
dead at the time of examination. Students
recorded that the SNAP Giardia fecal antigen
test identified 11 samples as Giardia positive
even though they did not identify any cysts
with the ZnSO4centrifugation technique
(Table 3). There was only one fecal sample that
was positivein the ZnSO4centrifugation tech-
nique and negative on the SNAP Giardia fecal
antigen test.
Twenty-seven participants returned complet-
ed forms from the Central Veterinary Confer-
ence wet lab. The centrifugation versus passive
flotation technique data from the wet lab
demonstrated that centrifugation with either
1.18-SG ZnSO4or 1.27-SG Sheather’s sugar
solution routinely recovered more eggs and
oocysts than the passive Ovassay technique
(Table 4). Not only did the centrifugation tech-
nique recover more eggs and oocysts, but the
participants also recorded many more samples
as positive with the centrifugation technique.
Only once did the Ovassay technique recover
all parasites in all samples; in contrast, only
once did the centrifugation technique fail to re-
cover all parasites in all samples (Table 4). Only
two of 14 participants in Group 1 (1.18-SG
ZnSO4solution) recovered T
aenia eggs using
the centrifugation procedure, whereas all 13
TABLE 2. Evaluation of Repeated 1.18-SG ZnSO4Centrifugation Fecal Flotations and
SNAP Giardia Tests to Identify Giardia-Positive Beagles: Trial 2, 2005
Pen No. of Week 1 Week 2 Week 3 Week 4 Week 5
No. of dogs or 4 6 6 6 6 6 6 7 7 7
pens testing
Total no. of 6 6 7 7 7
dogs or pens
testing positive
M. W. Dryden, P. A. Payne, and V. Smith
Veterinary Therapeutics Vol. 7, No. 1, Spring 2006
participants in Group 2 (1.27-SG Sheathers
sugar solution) recovered Taenia eggs using the
centrifugation technique (Table 4).
Even though the participants in the wet lab
weretold the samples were positive for Giardia
cysts, recoveryand identification of Giardia
cysts was problematic for the participants re-
gardless of the technique used. Only six of 27
participants wereable to recover and identify
Giardia cysts from a known positive sample:
One participant using the centrifugation tech-
nique with ZnSO4,one using Ovassay with
ZnSO4, one using Ovassay with Sheather’s sug-
ar solution, and three using the centrifugation
technique with Sheather’s sugar solution. All
27 participants had a positive SNAP Giardia
fecal antigen test on the sample provided.
A1.18-SG ZnSO4flotation with centrifuga-
tion should be adequate for the diagnosis of
Giardia spp bytrained personnel,2–4 but be-
cause many clinics do not use centrifugation
techniques and cyst shedding is intermittent, it
is often necessaryto examine several sequential
daily samples to ensureaccuracy of diagnosis.
The intermittent natureof cyst shedding was
particularly evident in the evaluation of sam-
ples from beagles during 2004. Inthe trials
evaluating repeated fecal examinations from re-
search beagles, a single ZnSO4fecal flotation
with centrifugation was positive 37.5% to
87.5% of the time in the 2004 investigation
and 57.14% to 100% of the time in the 2005
investigation. The sensitivity of the SNAP Gi-
ardia fecal antigen test was comparable, with
positive results obtained 50% to 62.5% of the
time in the 2004 investigation and 85.57% to
100% in the 2005 investigation. This further
demonstrates that a single negative ZnSO4fe-
TABLE 3. Evaluation by Second-Year Veterinary Students of 1.18-SG ZnSO4
Centrifugation Fecal Flotations and SNAP Giardia Tests to Identify Giardia spp in
SNAP+/ ZnSO4+/ SNAP+/ SNAP– / Combined
Category SNAP+ ZnSO4+ ZnSO4– SNAP– ZnSO4+ ZnSO4– Methods
No. of samples in 55 45 11 1 44 60 56
each category (total
no. of samples
tested = 116)
Percentage of 47.41% 38.79% 9.48% 0.86% 37.93% 51.72% 48.28%
samples in each
Percentage of 56 98.21% 80.36% 19.64% 1.79% 78.57%
samples recorded
as Giardia positive
identified as positive
by each specific
cal flotation with centrifugation or a negative
in-clinic Giardia fecal antigen test does not
necessarily rule out Giardia infection. Interest-
ingly, if three consecutive weekly ZnSO4fecal
flotations with centrifugation were performed
on the beagles in either trial, a positive diagno-
sis would haveoccurred in 100% of the bea-
gles. This is in agreement with a previous in-
vestigation that demonstrated that it may take
three consecutiveZnSO4fecal flotations to
achieve 94% accuracy in positive Giardia diag-
nosis.2The problem is that in a private prac-
tice, conducting three fecal examinations on
samples collected on consecutive days is often
Although the ZnSO4centrifugation proce-
dure conducted by a well-trained technician has
been considered the gold standard for the diag-
nosis of Giardia spp in dogs2–5 and was highly
accurate in the evaluation of the beagles when
the fecal samples were examined by a highly
trained technician, the second-year veterinary
students with less experience and training
recorded 11 samples as cyst negative that were
positiveon the SNAP Giardia fecal antigen test
kit. We werethe instructors during that class
TABLE 4. Veterinarian and Veterinary Technician Comparison of Ovassay and
Centrifugation Techniques for Recovery of Parasite Eggs and Oocysts Using 1.18-SG
ZnSO4aor 1.27-SG Sheather’s Sugar Solution
GROUP 1—ZnSO4(n = 14)
Ovassay Centrifugation
PARASITES 0 1–10 11–50 >50 % Positive 0 1–10 11–50 >50 % Positive
T. canis 0 7 7 0 100.0 0 1 4 9 100.0
T. vulpis 2 9 3 0 85.71 0 2 8 4 100.0
A. caninum 1 12 1 0 92.86 0 4 7 4 100.0
Taenia spp 13 1 0 0 7.69 12 2 0 0 14.29
Eucoleus boehmi 10 4 0 0 28.57 0 9 5 0 100.0
Cystoisospora spp 10 4 0 0 28.57 0 6 8 0 100.0
GROUP 2—Sheather’s Sugar Solution (n = 13)
Ovassay Centrifugation
PARASITES 0 1–10 11–50 >50 % Positive 0 1–10 11–50 >50 % Positive
T. canis 3 5 5 0 76.92 0 0 4 9 100
T. vulpis 2 7 4 0 84.62 0 0 6 7 100
A. caninum 7 4 2 0 46.15 0 0 7 6 100
Taenia spp 5 8 0 0 61.54 0 3 10 0 100
Eucoleus boehmi 10 3 0 0 23.08 0 4 9 0 100
Cystoisospora spp 7 6 0 0 46.15 0 3 10 0 100
aParticipants recorded the number of eggs recovered as 0, 1–10, 11–50, or >50 eggs/slide.
M. W. Dryden, P. A. Payne, and V. Smith
Veterinary Therapeutics Vol. 7, No. 1, Spring 2006
period and noted that while some of the sam-
ples contained numerous cysts (Figure 2), oth-
ers contained only a few cysts. We were unable
to examine every sample examined by the stu-
dents, but there were several samples that the
students identified as SNAP positive–float neg-
ative in which an instructor subsequently iden-
tified cysts. It must be noted that, in these in-
vestigations, we were unable to ascertain if any
of the SNAP positive–float negative samples
were in fact true false-positives.
The inability to recognize a few Giardia
cysts is likely one of the problems encountered
by the veterinarians and veterinary technicians
who participated in the wet lab. The sample
did not contain numerous Giardia cysts, and
they are smaller than the other parasite eggs
and oocysts in the sample. As wewalked
around the room to assist the participants, we
observed cysts in almost every sample evaluat-
ed using either ZnSO4or Sheather’s sugar so-
lution. Many of the participants were unable
to recognize the small cysts.
Since cyst shedding is notoriously intermit-
tent, the number of Giardia cysts recovered on
any given day is likely not a good indicator of
the level of infection. In a previous investiga-
tion, it was noted that dogs could have almost
a10-fold change in the number of cysts recov-
ered on quantitative fecal examinations con-
ducted 3 days apart.3Inthat study,the cysts/g
of one control dog went from less than 10 to
3,190 within 3 days. Inanother dog, the cyst/g
count increased from 150 to 44,610 within 3
days. The clinical implication of finding only a
fewGiardia cysts on fecal analysis may be no
different from finding hundreds. Therefore, fe-
cal examination from dogs or cats suspected of
having giardiasis requires careful microscopic
examination in case they are shedding low
numbers of cysts.
As noted previously, ZnSO4has been shown
to be the most efficient flotation solution for
recovery of Giardia cysts and is often used in
veterinary practices. The wet lab conducted at
the Central Veterinary Conference highlighted
apotential problem in using 1.18-SG ZnSO4,
even in a centrifugation procedure. Only two
of 14 (14.29%) participants using the 1.18-SG
ZnSO4centrifugation procedure correctly
recorded that the sample was positive for Tae-
nia eggs, while 100% of the participants using
the 1.27-SG Sheather’s sugar solution recov-
ered Taenia eggs from the same sample. This
result was not completely unexpected since
Taenia eggs have an average SG of 1.2251.7
This indicates that veterinary practices using
1.18-SG ZnSO4as their flotation solution are
likely failing to identify some dogs infected
with Taenia tapeworms and possibly other par-
asites that shed heavy eggs, such as Physaloptera
spp, which have eggs with an average SG of
1.2376.7Another investigation that evaluated
the SG of a fecal flotation solution indicated
that solutions with SGs of 1.22 to 1.35 would
be best for routine laboratory use.8
If giardiasis is on the differential list of a dog
(or cat) with diarrhea, the data indicate that
conducting both ZnSO4centrifugation fecal
flotation and a SNAP Giardia fecal antigen test
may increase the chances of recording a posi-
tive finding. However, it must also be remem-
bered that a single negative examination, even
if both tests areconducted simultaneously,
It may take three consecutive ZnSO4fecal flotations to
achieve 94% accuracy in positive Giardia diagnosis.
does not necessarily rule out giardiasis. Three
fecal examinations over a 7-day period using
both techniques will almost ensure a correct di-
agnosis. While using the proper Giardia cyst
recovery technique is important, identification
of recovered cysts is critical. In the Central Vet-
erinary Conference wet lab, registered veteri-
nary technicians and veterinarians had great
difficulty identifying cysts even when informed
the samples were positive. Proper training of
veterinarians and clinical staff on how to cor-
rectly identify Giardia cysts is important and
would greatly improve diagnostic accuracy, but
training or retraining of personnel already in
practices may be difficult on a large-scale basis.
The major question is, “What procedure or
procedures should be conducted for routine fe-
cal examinations?” Data from this current
study and another previously published study1
would suggest that the swing-head centrifuga-
tion technique using 1.27-SG Sheather’s sugar
solution is an efficient method of recovering
many commonly encountered parasite eggs
and oocysts. However, while the sugar solution
is effective for many eggs and oocysts, it dis-
torts and/or destroys most Giardia cysts, often
rendering them unrecognizable to most veteri-
narians and technicians (Figure 3). In addition,
currently used sugar solutions must be mixed
on site (see boxon page 7) and some chemical
needs to be added (e.g., phenol, formalin) to
prevent bacterial growth. Sugar solutions are
also sticky and can attract flies. Many practices,
therefore, have used commercially available salt
solutions for routine fecal examinations, in-
cluding ZnSO4. However,as demonstrated in
this investigation, a 1.18-SG ZnSO4flotation
solution may not be able to float parasite eggs
with a higher SG.
Because of the inability of 1.18-SG ZnSO4
flotation solution to consistently recover heav-
ier parasite eggs, it may be prudent for many
veterinary practices to conduct routine fecal
examinations using 1.27-SG Sheather’s sugar
solution. While some salt solutions may be
used as a practical alternative, the use of a cen-
trifugation technique will undoubtedly in-
crease diagnostic efficacy. Veterinary practices
should also consider the routine use of a hy-
drometer so that the proper SG of their flota-
tion solution can be assured. In addition, if gi-
ardiasis is encountered in the practice area,
fecal examinations should include a SNAP
Giardia fecal antigen test. The difficulty we
noted in the ability of veterinarians, veterinary
technicians, and veterinarystudents to identify
Giardia cysts in our studies is likely reflective
of the situation in many practices. Therefore,
the in-clinic soluble SNAP Giardia fecal anti-
gen Test Kit likely will improve a clinic’s abili-
ty to arrive at a correct diagnosis. In addition,
the proper recovery and identification of para-
sites should allowfor a moretargeted thera-
peutic approach.
We thank IDEXX Laboratories for partial support of this
study and for supplying the SNAP Giardia Test Kits.
Thanks also to the second-year veterinary students at
Kansas State University who took Veterinary Parasitology
during the fall of 2005. Wealso thank Bayer Animal
The in-clinic soluble SNAP Giardia fecal
antigen Test Kit likely will improve a clinic’s
ability to arriveat a correct diagnosis.
M. W. Dryden, P. A. Payne, and V. Smith
Veterinary Therapeutics Vol. 7, No. 1, Spring 2006
Health for sponsoring the wet lab at the Central Veterinary
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... Two grams of each fecal sample were processed using the flotation technique with zinc sulfate, as described by Dryden et al. [39]. A drop of fecal suspension was transferred to a microscope slide with a cover slip and examined at 40X magnification for identification of cysts. ...
... The diagnostic method used in the present study was zinc sulfate with centrifugation, which has proven to have a sensitivity of over 72% if a single sample is analyzed [48,49]. This technique has been considered the gold standard for the diagnosis of Giardia in dogs and cats [39]. However, due to the intermittent excretion of cysts and because only one sample was collected rather than the three recommended [49] the occurrence obtained in this study could be underestimated. ...
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Giardia is a protozoan that affects humans as well as a wide range of domestic species. It is distributed worldwide, and the highest frequency is seen in developing countries. Due to the potential for domestic cats to be carriers of this parasite and subsequently transmit the infection to humans, it is important to know the risk of transmission. For this reason, the objective of this study was to determine the frequency of this parasite in the cat population of the city of Santiago de Queretaro, Mexico, and identify the assemblages present to determine the role this host plays in public health, this being the first study of its type to be performed in the country. This was a cross-sectional study during which 200 fecal samples were collected from cats of both sexes and varying ages and strata of origin. The samples were analyzed by microscopy following the flotation technique, having obtained a general frequency of 25%. Giardia cysts were found at higher frequency in pasty stools. The assemblages found were zoonotic, specifically assemblage A, which suggests that the cat poses an important risk for the dissemination of the parasite to humans, making it an important public health problem.
... Misdiagnoses for giardiasis have been common and the actual prevalence may be underestimated [24]. It has also been reported that diagnosis of giardia infections using fecal smear is considered difficult because the cysts are small and similar in appearance to many pseudoparasites such as yeasts [25]. According to Kapoor [26], many patients who are treated for amoebiasis are actually suffering from giardiasis. ...
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The study was designed to investigate on selected practices among rural population, and their likely contribution to the spread of amoebiasis (E. histolytica) and giardiasis (G. lamblia). A cross sectional study was carried out in three villages, namely Kikapu, Piave and Belbar in Njoro District, Kenya. Questionnaire, interviews and personal observations were used to obtain data from 336 randomly selected consenting individuals in homesteads in the three villages. A retrospective study was further carried out to establish the prevalence of E. histolytica and G. lamblia among outpatients attending two randomly selected health centers in the study area. Majority of the respondents' highest education level was basic primary school education (61%), unemployment (98%) and with a monthly expenditure of less than 2200 Kenya Shillings or 28 US Dollars (81%). Eighty percent of the respondents were classified under low economic status. Eighty two percent reported lack of piped water and boiling of drinking water was less likely to occur among the low economic status respondents (Odds ratio (OR) = 0.423, 2 = 9.88; 95% CI of-5.74 to 6.58). However, washing of hands with soap after using a latrine seemingly was not influenced by economic status of the respondents (OR = 1; 2 = 0; 95% CI = 0). The level of education seemed to influence on the adoption of risky practices, such as, failure to boil drinking water was more likely to occur among respondents who had a low academic level (OR = 0.84, 2 = 0.04, 95% CI of-2.27 to 3.95). The stool tests records at Njoro PCEA health center showed that the prevalence of E. histolytica (20.83%) and G. lamblia (20.32%) were higher than the corresponding prevalence at Njoro County Council health center which were 1.34% and 0.00% respectively. Concurrent infections of E. histolytica and G. lamblia were absent in the two health centers. The trend of E. histolytica and G. lamblia followed an alternating pattern, in which an increase in one directly corresponded to a decrease in the other and vice versa in the two health centers. It was concluded that poverty and low education levels were significant factors that influenced on the adoption of risky lifestyles that were likely to enhance parasitic infections. There is a need for reliable diagnostic methods other than direct microscopy for E. histolytica and G. lamblia stool tests in order to minimize the wide variation of the results in the two health centers. Public health education should also be enhanced to discourage the adoption of risky practices.
... The use of chemical disinfectants, including chlorine and ozone, is limited by the production of toxic byproducts from reactions with compounds in the water. As G. lamblia trophozoites and cysts are widespread in the environment, untreated water from lakes, rivers, springs or shallow wells should not be consumed (Dryden et al. 2006). ...
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Giardia lamblia is the most common gastrointestinal disease causing agent in developed and developing countries. It has been reported that more than 280 million people are affected by acute and chronic giardiasis annually worldwide. The outcomes of G. lamblia infections are highly variable because of the mixed infections with other microorganisms. Majority of infected individuals are below 5 years. They are usually suffering from stomach cramp, irritable bowel syndrome, nausea and vomiting. Specific determinants for these diverse disease outcomes still remain elusive. Proper identification and genetic characterization of clinical isolates from endemic areas throughout the world becomes a certainly efficient tool to determine the potential genetic traits of parasite affecting its virulence capacity as well as different disease causing abilities. From the previous molecular biology studies, in both humans and animals, it is clear that predominance of the zoonotic Assemblage E subtype family of Giardia highlights the likely occurrence of zoonotic transmission in Indian cattle of giardiasis in West Bengal, Kolkata. This communication explores the fact that gastrointestinal diseases are the most numerous in humans because Giardia cyst is constantly exposed to the environment. The chapter throws light on protozoan diseases and highlights that low personal hygiene and contacts with animals are important predictors for intestinal protozoan infections. As it will emerge out from this communication, both anthroponotic and zoonotic transmissions play potential roles in the transmission of giardiasis in the community. It would also be clear as to how the molecular genotyping may assist in developing effective control strategies based on a better understanding of epidemiology of this parasite.
... Centrifugal flotation relies upon a centrifuge to separate particulate material based upon differential densities so that less dense ova or cysts float to the surface of the solution [22]. There are two types of centrifuges that you might choose to support fecal analysis in private practice: Regardless of which model you purchase, centrifugal flotation is considered the gold standard both in the clinic setting and at reference laboratories [4,7,8,[22][23][24][25][26]. Centrifugal flotation offers greater recovery of ova [8] as well as greater reliability in the diagnosis of Trichuris vulpis and Giardia lamblia [25], whereas passive fecal flotation may miss as many as 50.5% of infected dogs [27]. ...
The preceding chapter (Chapter 17) introduced direct smears or wet mounts as a convenient and expedient in‐house diagnostic tool that evaluates feces for motile organisms, including trophozoites. This chapter will explore an additional means by which we can microscopically examine feces. We will learn how to perform both passive (gravitational) and centrifugal flotation to recover cysts, oocysts, and eggs of mature parasites that live inside the body and reproduce. This technique relies upon differential densities to separate macerated feces within hypertonic solutions of concentrated sugar or salts.
... As it is often challenging to distinguish cysts from these contaminants, Giardia is considered one of the most "misdiagnosed, undiagnosed, or over-diagnosed parasites." 7 The most common method of Giardia identification is microscopy; however, it is often inaccurate and prone to human error. 6 Methods of purifying Giardia from fecal samples include density gradient centrifugation, 1 membrane-based filtration, gravity sedimentation, and flocculation. ...
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Giardia is one of the most common waterborne pathogens causing around 200 × 10 ⁶ diarrheal infections annually. It is of great interest to microbiological research as it is among the oldest known eukaryotic cells. Purifying Giardia from fecal samples for both research and diagnostic purposes presents one of the most difficult challenges. Traditional purification methods rely on density gradient centrifugation, membrane-based filtration, and sedimentation methods, which suffer from low recovery rates, high costs, and poor efficiency. Here, we report on the use of microfluidics to purify Giardia cysts from mouse feces. We propose a rigid spiral microfluidic device with a trapezoidal cross section to effectively separate Giardia from surrounding debris. Our characterizations reveal that the recovery rate is concentration-dependent, and our proposed device can achieve recovery rates as high as 75% with 0.75 ml/min throughput. Moreover, this device can purify Giardia from extremely turbid samples to a level where cysts are visually distinguishable with just one round of purification. This highly scalable and versatile 3D printed microfluidic device is then capable of further purifying or enhancing the recovery rate of the samples by recirculation. This device also has the potential to purify other gastrointestinal pathogens of similar size, and throughput can be significantly increased by parallelization.
Information concerning risk factors associated with Giardia infection in dogs in southern Ontario, Canada, is currently lacking. This study therefore aimed to identify risk factors for Giardia infection in dogs that visit off-leash dog parks in southern Ontario. From May-November 2018, fecal samples were collected from 466 dogs in 12 off-leash dog parks in the Niagara and Hamilton regions of Ontario. A survey that asked questions pertaining to travel history (i.e., area of residence, locations and regions visited in the previous 6 months), basic medical history (i.e., spay/neuter status, veterinary visits, use of deworming medication), consumption of a raw diet, and the physical (i.e., age, sex, breed) and behavioral characteristics (i.e., off-leash activities, hunting activities) of each dog sampled was administered to the respective owner. All fecal samples were examined with the Giardia plate ELISA (IDEXX Laboratories) for parasite antigen. Multivariable logistic regression analyses were conducted on the survey data to investigate putative risk factors for Giardia infection. Overall, 11.8% (95% CI: 9.2-15.1%) of samples tested positive for Giardia antigen. Results from the multivariable logistic regression analyses identified an interaction between dog age and spay/neuter status that was significantly associated with Giardia infection. The odds of infection were greater in intact as compared to neutered adult dogs (OR: 3.6, 95% CI: 1.7-7.9, p = 0.001), and in neutered juvenile dogs as compared to neutered adults (OR: 5.2, 95% CI: 2.2-12.2, p < 0.001). The results provide veterinarians with evidence-based information for identifying dogs at greatest risk of Giardia infection in southern Ontario.
The preceding chapter (Chapter 16) examined the physical properties of feces that can be appreciated by applying sight and smell to diagnostic medicine: fecal color, consistency, and odor. This chapter will expand our understanding of fecal diagnostics by transitioning from macroscopic to microscopic assessment. We will concentrate on how to perform direct smears or wet mounts. This technique is a convenient and expedient in‐house diagnostic tool that evaluates feces for motile organisms, including trophozoites, that may not survive the hypertonic solutions that are used in fecal flotation. For additional information of fecal flotation, please reference the next chapter, Chapter 18.
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Giardiosis is a worldwide intestinal parasitosis, affecting both humans and animals. Treatment in dogs remains limited and the lack of efficacy of the few approved medications is a rising concern. In this study, 23 dogs raised by veterinary students and naturally infected with Giardia duodenalis were treated in home conditions with fenbendazole (50 mg/kg orally for 5 consecutive days). Fecal samples were collected immediately before treatment (FS1), 2–4 days after treatment (FS2) and 8–10 days after treatment (FS3). G. duodenalis cyst excretion was measured quantitatively by direct immunofluorescence assay (DFA) at FS1, FS2 and FS3. Molecular typing with a nested PCR targeting the SSU rDNA locus was also performed at FS1 and FS2. Fecal consistency improved in 16/21 (76%) dogs and mean cyst shedding was reduced by 84% after treatment. However, only 8/23 (35%) dogs achieved therapeutic success (≥ 90% reduction of cysts) and only 4/23 (17%) dogs had complete elimination of G. duodenalis. Molecular typing showed that dogs harbored only canine-specific assemblages with a high prevalence of assemblage C in analyzed samples (30/39). We also detected different assemblages after treatment and nucleotide substitutions in assemblage C sequences that have not been previously described. Eight to ten days after treatment, high Giardia cyst excretion was measured, suggesting possible reinfection despite hygiene measures. These data suggest that fenbendazole treatment may improve fecal consistency but has limited therapeutic efficacy against giardiosis in this population of dogs. Further research is still needed to assess the efficacy of fenbendazole against canine giardiosis.
Fecal cytopathology is a component of a thorough diagnostic evaluation of patients with gastrointestinal (GI) signs and is useful for the identification of certain infectious agents as well as inflammatory and neoplastic cells. The collection method used affects the components retrieved, and rectal scraping is considered ideal for assessment of mucosal pathology. This chapter reviews the different methods of collection and the expected findings in healthy dogs and cats along with the incidental components of fecal evaluation. It also describes potential alterations in flora, inflammatory infiltrates, abnormal digesta, neoplasia, pathogens, and parasites, among others. Although fecal cytopathology most commonly aids in ruling out of selected causes of diarrhea, it occasionally provides a definitive diagnosis and should be a considered test modality for dogs and cats presented for GI disease.
Objective —To determine efficacy of treatment with a combination febantel-praziquantel-pyrantel product, with or without vaccination with a commercial Giardia vaccine, in dogs with naturally occurring giardiasis. Design —Prospective trial. Animals —16 Beagles naturally infected with Giardia duodenalis . Procedures —During phase 1, 6 dogs were treated with the parasiticide for 3 days (4 were also vaccinated). Four weeks later, all 6 dogs were treated with the parasiticide again for 5 days and were bathed and moved to clean cages after the last treatment (phase 2). Nine dogs were treated with the parasiticide for 3 (n = 4) or 5 (5) days and bathed and moved to clean cages after the last treatment (phase 3). Fecal samples were collected twice weekly for 24 days after treatment and tested for cysts with a quantitative zinc sulfate flotation technique and for Giardia antigen with an immunoassay. Results —Dogs in phase 1 were all shedding cysts again by day 24. In phase 2, only 1 dog shed cysts after treatment, and shedding was transient (day 17). In phase 3, neither cysts nor antigen was detected in fecal samples from 2 of 4 dogs treated for 3 days and 4 of 5 dogs treated for 5 days. In 18 of 57 (31.6%) fecal samples, cysts were seen, but results of the immunoassay were negative. Conclusions and Clinical Relevance —Results suggest that when a combination febantel-praziquantelpyrantel product is used to treat dogs with giardiasis, bathing and changing the environment after treatment may be more important in preventing recurrence than duration of treatment. ( J Am Vet Med Assoc 2002;220:330–333)
Two commercially available tests, an antigen-capture ELISA for use on fecal samples, and a peroral nylon string test for use in dogs, were compared with a zinc sulfate fecal concentration technique (ZSCT) for detection of giardiasis in dogs. Of 77 dogs and 164 fecal samples (from these dogs), 33 and 52, respectively were found to be Giardia-positive on the basis of results of the ZSCT. The ELISA gave false-negative results for 10 and 14% of ZSCT-positive dogs and fecal samples, respectively, and false-positive results (relative to the ZSCT test results) in 13 and 10% of ZSCT-positive dogs and fecal samples, respectively. Of the 18 string-tested dogs, 14 were positive by results of the ZSCT. Of the 4 dogs that were Giardia-negative by ZSCT, 2 were Giardia-positive by ELISA. Dogs were sedated and given water and metoclopramide to aid passage into the duodenum of the capsule containing a nylon string. Of the 21 string tests performed on the 18 dogs, only 5 strings reached the duodenum, and 0 of the 5 yielded positive results for Giardia sp. Because the string broke in 1 dog (leaving most in the gastrointestinal tract and, therefore, producing a risk of string foreign body) further string tests were not done.
The specific gravities of ten species of helminth eggs were determined using sucrose density gradient centrifugation. Fecal or egg concentrate was layered over a 3 to 54% sucrose density gradient. The gradient was then centrifuged at 800 g for 20 min, allowing 5 min for acceleration and 5 for deceleration. Bands formed were identified and measured. Refractive index was measured at the middle of narrow bands, or at the level at which the concentration of eggs was highest, in the case of wide bands or when no band was formed. The specific gravity corresponding to this refractive index was taken as the specific gravity of the eggs. The ten species of helminth eggs studied and specific gravities measured on three or four gradients were: Toxascaris leonina, 1.0559; Ancylostoma caninum, 1.0559; Toxocara canis, 1.0900; Parascaris equorum, 1.0969; Toxocara cati (embryonated), 1.1005; Ascaris suum, 1.1299; Trichuris suis, 1.1299; Trichuris vulpis, 1.1453; Taenia sp., 1.2251; and Physaloptera sp., 1.2376. These determinations agree with or approximate those of previous workers. The specific gravities of P. equorum, T. suis, Taenia sp., and Physaloptera sp., are reported for the first time.
Several variables in a standard vial fecal gravitational flotation technique were investigated. These were the specific gravity of the sodium nitrate flotation solution, duration of flotation and mesh sizes of strainers. The number of eggs which floated and adhered to a coverslip were counted and estimates of the number of eggs remaining in the strained fecal suspension and in the feces trapped on the strainer were made. Eggs from hookworms, Trichuris vulpis and Toxocara canis in feces from dogs, Nematodirus spp. from sheep and Parascaris equorum from horses floated equally well in solutions with specific gravities (SpGr) ranging from 1.22-1.38. Taenia spp. from dogs had a slightly narrower range (SpGr 1.27-1.38) for best recovery. Eggs from Haemonchus contortus from sheep appeared to float best between SpGr 1.22- 1.32. Strongyles from one horse floated best with SpGr 1.27-1.32 and from another with SpGr 1.11-1.38. Coccidial oocysts from sheep floated best in a narrow range of SpGr from 1.22-1.27. However, as the SpGr of the solution was increased the recognition of eggs under the coverslip was increasingly difficult and especially so at SpGr 1.38 with sheep feces. This was due to the increasing amount of debris and the more rapid formation of crystals with evaporation with solutions of higher SpGr. It appeared, therefore, that solutions with SpGr of 1.22-1.35 would be best for routine laboratory use. At specific gravity 1.27, there appeared to be no difference in the number of eggs recovered for a four, eight and 12 min flotation period. Only 3-7% of the eggs in 4 g of feces were counted under the coverslip. This poor efficacy resulted first because approximately 50% of the eggs were trapped in the feces and retained on the strainer. Secondly, only one half of the strained fecal suspension, containing approximately 25% of the eggs, was placed in the vial for examination. Thirdly, of those eggs in the vial only 16-29% were counted under the coverslip. When the second half of the strained fecal suspension was placed in another vial, the amount of debris and air bubbles adhering to the coverslip was much less than that for the first vial. Egg counts for both vials appeared similar and it may be that when debris is excessive the fecal examination should involve counts from a second vial. The use of strainers finer than the standard tea strainer and the addition of minimal amounts of detergent did not increase the egg count.
A capture enzyme linked immunosorbent assay (CELISA) was evaluated for its ability to detect Giardia coproantigens in the faeces of humans and dogs in the Perth metropolitan area and Aboriginal communities in Fitzroy Crossing, Western Australia. Using zinc sulphate flotation and light microscopy, Giardia cysts and/or trophozoites were observed in 8 of 57 (14%) human stool samples from Perth and 21 of 55 (38%) stool samples from Fitzroy Crossing, after 2 separate examinations. Analysis of diagnostic sensitivity using the ELISA revealed that coproantigens were detected in all 29 human samples (100%) in which Giardia cysts and/or trophozoites were also present. Coproantigens were detected in one further sample from Perth and in 3 samples from Fitzroy Crossing in which no Giardia cyst or trophozoite was observed. The specificity of the test, as defined using Fitzroy Crossing samples free from Giardia, was 91%. The assay did not cross-react with Giardia-free stool samples containing Hymenolepis nana, Entamoeba coli, E. hartmanni, Chilomastix mesnili or Ancylostoma duodenale. Giardia cysts and/or trophozoites were also observed in 11 of 32 dog faecal samples (34%) in Perth and 11 of 29 dog samples (38%) in Fitzroy Crossing, after one zinc sulphate examination. The sensitivity of the ELISA for dogs was 64% and 55% for Perth and Fitzroy Crossing specimens respectively. The specificity was 95% when Fitzroy Crossing samples were used. Other parasites observed in Giardia-free faecal samples from dogs which did not produce a positive reaction with the kit were Ancylostoma caninum, Sarcocystis sp. and Isospora sp.(ABSTRACT TRUNCATED AT 250 WORDS)
Fifty canine fecal samples were evaluated by five flotation procedures to compare the sensitivity of the zinc sulfate (ZnSO4) centrifugation flotation test with ZnSO4 flotation tests using benchtop incubation during the flotation period. One or more parasite species were detected in 40 samples. Results showed that centrifugation with ZnSO4 solution was significantly more likely to detect a positive sample than benchtop procedures. The difference in procedures was due primarily to increased detection of Trichuris eggs and Giardia cysts by centrifugal flotation. No significant difference was seen in the ability of benchtop procedures to detect positive samples when tests sat either for 5 or 10 minutes before examination.
A variety of procedures are available to detect parasite eggs or oocysts in feces. This study compared the efficacy of simple flotation, a commercial assay, and various centrifugation techniques and three common flotation solutions. Results indicate that centrifugation consistently recovered more eggs than other methods. Proper technique is critical, including ensuring that the specific gravity of the flotation solution is correct and allowing the sample to stand for a sufficient amount of time before examining the coverslip. Because of the zoonotic health risks of many companion animal parasites, veterinarian and their staff should better utilize fecal examinations in their routine diagnostic plan.
Evaluation of the efficacy of Drontal Plus and GiardiaVax to elimi-nate cyst shedding in dogs naturally infected with Giar-dia sp
  • P Payne
  • M Dryden
  • R Ridley
Payne P, Dryden M, Ridley R, et al: Evaluation of the efficacy of Drontal Plus and GiardiaVax to elimi-nate cyst shedding in dogs naturally infected with Giar-dia sp. JAVMA 220(3):330–333, 2002
Evaluation of the efficacy of Drontal™ Plus and GiardiaVax™ to eliminate cyst shedding in dogs naturally infected with Giardia sp
  • P Payne
  • M Dryden
  • R Ridley
Payne P, Dryden M, Ridley R, et al: Evaluation of the efficacy of Drontal™ Plus and GiardiaVax™ to eliminate cyst shedding in dogs naturally infected with Giardia sp. JAVMA 220(3):330-333, 2002.