CHAPTER 2.1 (OTHER SPECIMENS AND TECHNIQUES) Flashcards
other parasite detection techniques aside from the usual O&P methods
These are often used when the
physician suspects a particular parasite and the traditional O&P examination is negative.
These procedures include examination of?
duodenal material, sigmoidoscopy material,
and using cellophane tape to detect pinworms
Parasites that reside in the small intestine may be more difficult to recover in a stool specimen. In these situations, examining material from the ______ may yield success
duodenal area
How is duodenal material collected
Nasogastric Intubation or the Enteric Capsule Test (Enterotest)
Parasites that may be observed in duodenal material includes
Giardia intestinalis trophozoites, Cryptosporidiumspp.,
Isospora belli,
Strongyloides stercoralis,
eggs of Fasciola hepatica or Clonorchis sinensis
Duodenal fluid can be examined at a later time because even if there are trophozoites present, they will not deteriorate rapidly compared to normal fecal samples
true or false
False
Duodenal material can be examined
microscopically as a wet preparation. If the
volume of fluid is sufficient (>3 mL)
True or False
False >2 ml
In examining duodenal material, what type of concentration technique; fixative; and stain is used?
Sedimentation
PVA fixative
Trichrome/ Iron hematoxylin/ Acid-fast stain
Duodenal material can be used to perform antigen tests for
Cryptosporidium and/or Giardia
Explain the step by step process of Enterotest in collecting duodenal material
- Px SWALLOWS a gelatin capsule that contains a coiled length of yarn
- The capsule DISSOLVES
in the stomach and the weighted string is
carried to the duodenum. - The FREE END of the string is attached to the patient’s neck or cheek with tape
- After a 4-hour incubation period, the
yarn is PULLED BACK out of the patient - The bile stained mucous material brought up on the string is then EXAMINED microscopically via wet preps and, if necessary, permanent stains.
is often helpful for detecting E. histolytica.
Examination of sigmoidoscopy (colon) material
Material from ulcers obtained by aspiration or scraping should be examined by
direct wet preparations and permanent stains
Other species that may also be recovered from examining material from the sigmoid colon.
Coccidian parasites and microsporidia
If E. histolytica is present, it indicates what stage will also be often present?
trophozoite stage
What other material can be collected from the colon?
Colon biopsy material
Colon material are best processed using surgical pathology methods when these organisms are suspected to be present
amebae
is the specimen of choice for the detection of Enterobius vermicularis (pinworm) eggs; Adult female pinworms may also be seen
cellophane tape prep
Why is it important that the specimen be
collected in the morning before the patient washes or defecates for cellophane tape prep?
At night, when the body is at rest, pregnant adult female worms exit the host, typically a child, through the rectum and lay numerous eggs in the perianal region
In addition to pinworm, there is evidence to support the use of Cellophane tape prep technique for the recovery of
Taenia spp. eggs
the standard protocol for specimens collected daily for the number of negative tests that should be performed to rule out a pinworm infection is
Five
From which area can the Enterotest be used to collect specimens?
A. Duodenum
B. Sigmoid colon
C. Stomach
D. Perianal area
A. Duodenum
Systemic or blood-borne parasitic infections are diagnosed by demonstrating the
diagnostic stage(s) of the responsible parasite(s) in a blood specimen
Parasites that may be recovered in
blood include
Leishmania donovani spp
Trypanosoma spp
Plasmodium spp
Babesia spp.
microfilariae
can be detected by observing motility in a
wet preparation of a fresh blood sample under low- and high-power magnification
Trypanosoma spp.
microfilariae
Blood specimens for
parasite study must be collected by
aseptic technique
Although these specimens provide the best morphology of the parasites, improper collection or smear preparation can lead to unsatisfactory results
Blood from the fingertip or earlobe
Capillary blood collected for detecting parasites should be
- free-flowing and not contaminated with the alcohol
- Not milked if from the finger
Why shouldn’t we milk the fingers when collecting blood samples from the finger for detecting parasites
Diluted by tissue fluids making it difficult to detect parasites
Anticoagulants causes what to samples for parasite detection in blood?
distortion to the staining
process and subsequent parasite morphology
Blood specimens that is collected through venipuncture should be collected in tubes containing
ethylenediaminetetraacetic
acid (EDTA)
If malaria is suspected,
it is best to prepare smears within
1 hour
Stage in detecting parasites in blood which consists of preparing thick and thin
blood smears, staining them using a permanent stain, and examining them microscopically
blood sample processing
Blood samples may also be processed by performing the ________
which is characterized by examining buffy
coat slides, or setting up and reading cultures
Knott technique
are frequently satisfactory for screening purposes, particularly when malaria is suspected.
Thick smears
provide the best view of the malarial
parasites in red blood cells and are recommended for species identification.
Thin smears
What type of thick smears allows them to typically have a much higher concentration of parasites than thin smears
dehemoglobinized thick smears
Thick smears are primarily used when
parasites are few in number or when
thin smears are negative
The advantage of the thick smear is
increased ability to detect the
malarial parasites
The disadvantage of the thick smear is
red blood cells have been lysed and it is not
possible to assess the morphology of parasites that are detected
There are two permanent
stains commonly used for the detection of blood parasites namely?
Wright’s stain and Giemsa stain
Stain that contains the fixative
and stain in one solution,
Wright’s stain
Stain where the fixative
and stain are not in one solution,
Giemsa stain
Wright’s stain typically yields only
satisfactory results
is considered the preferred stain because it allows for the detection of parasite detail necessary for species identification
Giemsa stain
Appearance of Select
Parasitic Structures and
Background Material on
GIEMSA STAIN
RED
Leishmania, trypanosome,
malaria, and Babesia nuclear
structures
Appearance of Select
Parasitic Structures and
Background Material on
GIEMSA STAIN
BLUE
Cytoplasm
Appearance of Select
Parasitic Structures and
Background Material on
GIEMSA STAIN
also… RED
Schüffner’s dots
Appearance of Select
Parasitic Structures and
Background Material on
GIEMSA STAIN
BLUE TO PURPLE
Nuclei
Appearance of Select
Parasitic Structures and
Background Material on
GIEMSA STAIN
CLEAR; MAY NOT STAIN
Sheath
Appearance of Select
Parasitic Structures and
Background Material on
GIEMSA STAIN
PALE RED
Red blood cells
Appearance of Select
Parasitic Structures and
Background Material on
GIEMSA STAIN
PURPLE
White blood cells
Appearance of Select
Parasitic Structures and
Background Material on
GIEMSA STAIN
PINK-PURPLE
Neutrophilic granules
Appearance of Select
Parasitic Structures and
Background Material on
GIEMSA STAIN
PURPLE-RED
Eosinophilic granules
is designed to concentrate blood specimens suspected of containing low numbers of microfilariae.
Knott Technique
How to do a simple modified version of KNOTT TECHNIQUE
combining 1 mL of venipuncture collected
blood with 10 mL of 2% formalin in
a centrifuge tube thoroughly mixed and spun for 1 minute at 500 × g.
What can we collect from the resulting
sediment of KNOTT TECHNIQUE
Thick slides may be made, dried, and
subsequently Giemsa-stained
Buffy coat cells may be extracted
from blood specimens, stained with Giemsa
stain, and microscopically examined for
Leishmania and Trypanosoma
How to collect and make buffy coat slides
collecting oxalated or citrated blood,
placing it in a Wintrobe tube, and spinning it for 30 minutes at 100 × g.
buffy coat may then be extracted using a capillary pipette
One such culture technique
that yields favorable results for the recovery
of Leishmania spp. and Trypanosoma cruzi
uses
Novy-MacNeal-Nicolle (NNN) medium.
Novy-MacNeal-Nicolle (NNN) medium is used in culture techniques to detect what parasites
Leishmania spp. and Trypanosoma cruzi
give the step by step procedure of the NNN medium
- NNN slant is inoculated by the ADDITION of a single drop of collected blood or ground tissue
- Penicillin is added to the medium if the specimen originates from a source that may contain bacteria
- PERIODIC EXAMINATION, every other day, should be conducted by observing the slant under 400× magnification.
- Negative cultures should be held
for 1 month.
Cerebrospinal fluid (CSF) specimens may be collected for the diagnosis of
amebic conditions associated with select ameba
and African sleeping sickness
A wet preparation of CSF samples
can be prepared to search for the presence of the characteristic morphologic forms of
Naegleria fowleri and Acanthamoeba spp and the trypomastigote stages of Trypanosoma spp.
Special stains can also be performed on CSF including
Giemsa, trichrome, and modified trichrome
stains
If Naegleria or Acanthamoeba are suspected
of being potential pathogens, the CSF specimen can be cultured on
non-nutrient agar seeded with Escherichia coli
The CSF sediment is inoculated
to the medium, sealed, and incubated at what temperature
35° C
Other pathogens that might be recovered from the central nervous system include
Toxoplasma gondii and microsporidia and Taenia solium cysticercus larvae and Echinococcus spp.
Sterile fluids other than CSF include several
specimen types, such as
fluid present in cysts,
aspirates,
peritoneal fluid,
pleural fluid, and
bronchial washings
Tissue and biopsy specimens are recommended for the recovery of a number of parasites, including intracellular organisms such as
Leishmania spp. and T. gondii
is the preferred method for handling tissue and biopsy samples
Surgical removal of the specimen followed by the preparation of histologic tissue sections and impression smears
Other parasites that may be detected
in Tissue and Biopsy samples include
free-living ameba, Trypanosoma
spp., Trichinella spiralis, and microsporidia
is the specimen of choice for patients suspected of liver abscesses caused by E. histolytica
Hepatic abscess material
liver abscesses is caused by
Entamoeba histolytica
Sputum is typically collected and tested from
patients suspected of being infected by the
lung fluke: Paragonimus westermani
Patients with ________ will demonstrate motile larvae in their sputum
Strongyloides stercoralis hyperinfection
Other parasitic infections that
may be found in sputum samples include
microsporidia, E. histolytica, Entamoeba gingivalis, Ascaris lumbricoides, and hookworm
What type of specimen is recommended for sputum collection
early-morning specimen
The sputum sample may then be examined directly via
wet preps and/or concentrated using
N-acetylcysteine or other appropriate agent
Urine is the specimen of choice for the detection of
Schistosoma haematobium eggs
and Trichomonas vaginalis trophozoites
can sometimes be found in the urine of patients with a heavy filarial infection.
Microfilariae
The procedure for examining urine specimens for parasite detection
- Collected in a clean watertight container
- centrifuged on arrival
- Microscope analysis of sediment
are typically collected and examined for the presence of T. vaginalis trophozoites.
Vaginal and urethral specimens, as well as
prostatic secretions
are the method of choice for demonstrating the motile trophozoites in urine samples
Saline wet preparations
Alternative techniques for the diagnosis of
T. vaginalis include
antigen detection methods using latex agglutination and EIA procedures
commercially available nucleic acid probe
commercial product that uses a
culture pouch
is best diagnosed by the collection and examination of corneal scrapings
Acanthamoeba keratitis
Other specimens that may be tested for eye infections include a
contact lens or contact lens solution
The eye samples may be processed in several ways
the first way is?
- cultured on an agar plate seeded
with gram-negative bacteria.
Examining the culture plate from eye specimens should be done under
low dry magnification every day for 1 week
Examining the culture plate from eye specimens should reveal
the trophozoites >4 days
and the cysts in 4 to 5 days
The eye samples may be processed in several ways
the second way is?
the scrapings may be transferred
to glass slides and stained
Eye scrapings observed under the glass slides are stained using?
calcofluor white stain
Eye scrapings observed under the glass slides are microscopically observed using what type of microscope?
fluorescent microscopy
In eye specimen samples, what parasite stains apple green when calcoflour white stain is used?
Acanthamoeba cysts
Eye specimen samples when gone through staining procedures for parasite detection cannot stain what stage of the parasite?
trophozoites
In addition to Acanthamoeba, T. gondii, what are other potential eye pathogens
microsporidia, and Loa loa
microsporidia, and Loa loa can be detected using what?
histologic stains and specialized culture
methods
Mouth scrapings are the sample of choice for the detection of
E. gingivalis and Trichomonas tenax
nasal discharge specimens are helpful for the recovery of parasites such as
N. fowleri
Material obtained via mouth scrapings
and nasal discharge should be placed in a
Airtight collection container (swab or cup)
What techniques do we apply to mouth scrapings and nasal discharge?
Wet preps are more common
Permanent stains when applicable
Skin Snips are useful in the detection of
Onchocerca volvulus
Skin snips may be made using one
of two collection techniques. The objective of
both procedures is to obtain skin fluid without bleeding.
the first one is?
making a firm (scleral) punch into skin with a specially designed tool (sinearch ko: Gass Corneoscleral Punch maybe)
Skin snips may be made using one
of two collection techniques. The objective of
both procedures is to obtain skin fluid without bleeding.
the second one is?
uses a razor blade with which a small cut into the skin is made
Explain the step by step way we examine skin snips
- Tissues collected are placed in 0.2mL saline
- 30 minute incubation
- Microscopically examined
- Jerky movement of the microfilariae should be visible as they migrate from skin snip to the liquid saline
are not a common means of detecting parasites. There are a few techniques available but they are not usually performed in the routine laboratory. Specialized laboratories and research facilities may offer these services.
Culture methods
Parasites that can be isolated with culture include
E. histolytica
T. vaginalis
Leishmania spp.
T.cruzi
T. gondii
Animal Inoculation and Xenodiagnosis are the appropriate techniques to use on appropriate specimens from patients suspected of suffering from
Leishmania and Trypanosoma as well as Toxoplasma
Certain parasites have host specificity and require particular animals such as
Mice, guinea pigs, and hamsters
Leishmania and Trypanosoma as well as Toxoplasma if suspected, these techniques are used
Animal Inoculation and
Xenodiagnosis
is a technique that uses the arthropod host as an indicator of infection.
Xenodiagnosis
Suitable specimens for animal inoculation vary depending on the parasite suspected; these include
blood, lymph node aspirates, CSF, and
bone marrow
The specimens for animal inoculation and xenodiagnosis should be collected
using
aseptic technique
Xenodiagnosis is a technique used for the
diagnosis of what disease
Chagas’ disease
Explain how xenodiagnosis techniques is used for the diagnosis of Chaga’s disease
- An uninfected reduviid bug is allowed to take a blood meal from the patient
- bug’s feces is then examined to observe for the presence of T. cruzi
Xenodiagnosis to diagnose Chaga’s disease is primarily used in?
South America and Mexico.
Thick blood smears for malaria are recommended for
species identification
A. True
B. False
Giemsa is the preferred stain for the detection of blood parasites.
A. True
B. False
Which of the following is the specimen of choice to demonstrate intracellular parasites such as Toxoplasma gondii and Leishmania spp.?
A. Sputum
B. Urine
C. Tissue
D. Genital secretions
Is usually considered as an adjunct or supplement to standard laboratory protocols. Include methods for antigen
and antibody detection.
Immunologic tests
Occasionally, however, standard laboratory
tests are not sufficient for the diagnosis of
a parasite. For example, in some parasitic infections, the diagnostic stage is located deep in the tissues of the host
give an example of an infection like this
Toxoplasmosis
Why is standard laboratory test not sufficient for the diagnosis of toxoplasmosis
it may not be possible to detect
its presence
What disease can standard laboratory test not diagnose and may be dangerously invasive to attempt it
echinococcosis
As standard laboratory test are not sufficient for diagnosing toxoplasmosis and echinococcosis what is the alternative?
immunologic assays can be used
(IMMUNLOGIC TESTS)
The presence of an antibody
against a given parasite may not always indicate a current infection, WHY?
antibodies remain with a host for many years, a positive test result can occur from a past infection
What can we safely consider as a positive result from immunologic tests?
detection of an antibody to a given parasite in a patient with no previous exposure prior to travel to an endemic area
What org performs immunologic assays on
request
Centers for Disease Control and Prevention
(CDC)
(probably Research Institute for Tropical Medicine- National reference laboratory for parasites pag sa Philippines)
The only commercial molecular test available is for the diagnosis of
T. vaginalis
The detection of an antibody to a given parasite in a patient with no previous exposure prior to travel to an endemic area can be considered a positive result.
A. True
B. False
A. True
BF
Bentonite flocculation
CA
card agglutination
CF
complement fixation
DFA
direct fluorescent antibody
EIA
enzyme immunoassay
IB
immunoblot
IHA
indirect hemagglutination
IFA
indirect fluorescent antibody
LA
latex agglutination
PCR
polymerase chain reaction
Rapid
immunochromatographic cartridge
Cite the Parasitic Diseases that can be tested through Immunoassays and Molecular Techniques
African trypanosomiasis
Amebiasis
Babesiosis
Chagas’ disease
Cryptosporidiosis
Cysticercosis
Echinococcosis
Fascioliasis
Filariasis
Giardiasis
Leishmaniasis
Malaria
Microsporidiosis
Paragonimiasis
Schistosomiasis
Strongyloidiasis
Toxocariasis
Toxoplasmosis
Trichinellosis
Trichomoniasis
Cite the Immunoassays and Molecular Techniques for these Parasitic Diseases
African trypanosomiasis
Antigen Test: –
Antibody Test: CA, IFA
Molecular Test: PCR
Cite the Immunoassays and Molecular Techniques for these Parasitic Diseases
Amebiasis
Antigen Test: EIA, IFA
Antibody Test: EIA, IHA
Molecular Test: PCR
Cite the Immunoassays and Molecular Techniques for these Parasitic Diseases
Babesiosis
Antigen Test: –
Antibody Test: IFA
Molecular Test: PCR
Cite the Immunoassays and Molecular Techniques for these Parasitic Diseases
Chagas’ disease
Antigen Test: –
Antibody Test: CF, EIA, IFA
Molecular Test: PCR
Cite the Immunoassays and Molecular Techniques for these Parasitic Diseases
Cryptosporidiosis
Antigen Test: DFA, EIA, IFA, Rapid
Antibody Test: –
Molecular Test: PCR
Cite the Immunoassays and Molecular Techniques for these Parasitic Diseases
Cysticercosis
Antigen Test: –
Antibody Test: EIA, IB
Molecular Test: –
Cite the Immunoassays and Molecular Techniques for these Parasitic Diseases
Echinococcosis
Antigen Test: –
Antibody Test: EIA,IB
Molecular Test: –
Cite the Immunoassays and Molecular Techniques for these Parasitic Diseases
Fascioliasis
Antigen Test: –
Antibody Test: EIA,IB
Molecular Test: –
Cite the Immunoassays and Molecular Techniques for these Parasitic Diseases
Filariasis
Antigen Test: Rapid
Antibody Test: EIA
Molecular Test: –
Cite the Immunoassays and Molecular Techniques for these Parasitic Diseases
Giardiasis
Antigen Test: DFA, EIA, Rapid
Antibody Test: –
Molecular Test: PCR
Cite the Immunoassays and Molecular Techniques for these Parasitic Diseases
Leishmaniasis
Antigen Test: Rapid
Antibody Test: EIA, IFA
Molecular Test: PCR
Cite the Immunoassays and Molecular Techniques for these Parasitic Diseases
Malaria
Antigen Test: Rapid
Antibody Test: IFA
Molecular Test: PCR
Cite the Immunoassays and Molecular Techniques for these Parasitic Diseases
Microsporidiosis
Antigen Test: IFA
Antibody Test: –
Molecular Test: –
Cite the Immunoassays and Molecular Techniques for these Parasitic Diseases
Paragonimiasis
Antigen Test: –
Antibody Test: EIA, IB
Molecular Test: –
Cite the Immunoassays and Molecular Techniques for these Parasitic Diseases
Schistosomiasis
Antigen Test: EIA
Antibody Test: EIA, IB
Molecular Test: –
Cite the Immunoassays and Molecular Techniques for these Parasitic Diseases
Strongyloidiasis
Antigen Test: –
Antibody Test: EIA
Molecular Test: –
Cite the Immunoassays and Molecular Techniques for these Parasitic Diseases
Toxocariasis
Antigen Test: –
Antibody Test: EIA
Molecular Test: –
Cite the Immunoassays and Molecular Techniques for these Parasitic Diseases
Toxoplasmosis
Antigen Test: –
Antibody Test: EIA, IFA, LA
Molecular Test: PCR
Cite the Immunoassays and Molecular Techniques for these Parasitic Diseases
Trichinellosis
Antigen Test: –
Antibody Test: BF, EIA
Molecular Test: –
Cite the Immunoassays and Molecular Techniques for these Parasitic Diseases
Trichomoniasis
Antigen Test: DFA, LA, Rapid
Antibody Test: –
Molecular Test: DNA probe
