Microscopic Examination

Good, clean microscopes and light sources are mandatory when examining specimens for parasites. Organism identification depends on morphologic differences, most of which must be seen using stereoscopic (magnification s50x) or regular microscopes at low (lOOx), high dry (400x), and oil immersion (lOOOx) magnifications. The use of a 50x or 60x oil immersion objective for scanning can be very helpful, particularly when the 50x oil and lOOx oil immersion objectives are placed side by side,, This arrangement on the microscope can help avoid accidentally getting oil on the 40x high dry objective.

A stereoscopic microscope is recommended for larger specimens (e.g., arthropods, tapeworm proglotj tids, various artifacts). The total magnification usual^j varies from approximately 10x to 45x, either with a zoom capacity or with fixed objectives (0.66x, 1.3x, 3x) that can be used with 5x or 10x oculars. Depending on the density of the specimen or object being examine^ the light source must be directed either from under the stage or onto the top of the stage.

Intestinal Tract

Stool is the most common specimen submitted to the. diagnostic laboratory; the most commonly perform«^ procedure in parasitology is the ova and parasite (0&r| examination. Several other diagnostic techniques arej available for the recovery and identification of parasiffl organisms from the intestinal tract. Most laboratories d(j not routinely offer all of these techniques, but many «4J relatively simple and inexpensive to perform. The cM' cian should be aware of the possibilities and the din^3' relevance of information obtained from using sucft techniques. Rarely, it is necessary to examine stool spcj cimens for the presence of scolices and proglottid^ ot cestodes and adult nematodes and trematodes t0

PRINCIPLE:

To assess worm burden of patient, to provide quick diagnosis of heavily infected specimen, to check organism motility (primarily protozoan trophozoites), and to diagnose organisms that might not be seen from the permanent stain methods; seeing organism motility is the primary objective SPECIMEN:

Any fresh liquid or very soft stool specimen that has not been refrigerated or frozen

REAGENTS:

0.85% NaCI; Lugol's or D'Antoni's iodine EXAMINATION:

Low-power examination (x100) of entire 22- by 22-mm coverslip preparation (both saline and iodine); high dry power examination (x400) of at least one third of the coverslip area (both saline and iodine)

RESULTS AND LABORATORY REPORTS:

Hesults from the direct smear examination should often be considered presumptive; however, some organisms could be definitively identified {Giardia lambtia cysts and Entamoeba coli cysts, helminth eggs, and larvae, Isospora belli oocysts). These reports should be considered "preliminary," while the final report would be available after the results of the concentration and permanent stained smear were available PROCEDURE NOTES AND LIMITATIONS: Once iodine is added to the preparation, the organisms will be killed and motility will be lost. Specimens submitted in stool preservatives or fresh, formed specimens should not be examined using this procedure; the concentration and permanent stained smear should be performed instead. Oil Immersion examination (x1000) is not recommended (organism morphology not that clear)

confirm the diagnosis and/or for identification to the species level.

Other specimens from the intestinal tract, such as duodenal aspirates or drainage, mucus from the Entero-Test Capsule technique, and sigmoidoscopy material, can also be examined as wet preparations and as permanent stained smears after processing with either tri-chrome or iron-hematoxylin staining. Although not all laboratories perform these procedures, they are included to give some idea of the possibilities for diagnostic testing.

O&P EXAMINATION. The O&P examination comprises three separate protocols: the direct wet mount, the concentration, and the permanent stained smear.*

The direct wet mount requires fresh stool, is designed to allow detection of motile protozoan trophozoites, and is examined microscopically at low and high dry magnifications (xlOO, entire 22- x 22-mm Coverslip; x400, lh tolh 22- x 22-mm coverslip) (Box 49-1), However, because of potential problems resulting iiom the lag time between specimen passage and receipt the laboratory, the direct wet examination has been

BOX 49-2 Concentration—Review

PRINCIPLE:

To concentrate the parasites present, either through sedimentation or by flotation. The concentration is specifically designed to allow recovery of protozoan cysts, coccidian oocysts, microsporidian spores, and helminth eggs and larvae SPECIMEN:

Any stool specimen that is fresh or preserved in formalin (most common), PVA (mercury- or non-mercury-based), SAfi MIF, or the newer single-vial-system fixatives REAGENTS:

5% or 10% formalin, ethyl acetate, zinc sulfate (specific gravity, 1.18 for fresh stool and 1.20 for preserved stool); 0.85% NaCI; Lugol's or D'Antoni's iodine. EXAMINATION:

Low-power examination (x100) of entire 22- by 22-mm coverslip preparation (iodine recommended, but optional); high dry power examination (x400) of at least one third erf the coverslip area (both saline and iodine)

RESULTS AND LABORATORY REPORTS:

Often, results from the concentration examination should be considered presumptive; however, some organisms could be definitively identified (Giardia lamblla cysts and Entamoeba coli cysts, helminth eggs, and larvae, Isospora belli oocysts). These reports should be considered "preliminary," while the final report would be available after the results of the permanent stained smear were available

PROCEDURE NOTES AND LIMITATIONS:

Formalin-ethyl acetate sedimentation concentration is the most commonly used. Zinc sulfate flotation will not detect opereulated or heavy eggs (Clonorchls eggs, unfertilized Ascaris eggs); both the surface film and sediment will have to be examined before a negative result is reported. Smears prepared from concentrated stool are normally examined at low (x100) and high dry (x400) power; oil immersion examination (x1000) is not recommended (organism morphology not that clear). The addition of too much iodine may obscure helminth eggs (will mimic debris)

eliminated from the routine O&P in favor of receipt of specimens collected in stool preservatives. The direct wet preparation is not performed for specimens received in the laboratory in stool collection preservatives. Specific fixatives can be found in Table 49-8.

The second part of the O&P is the concentration, which is designed to facilitate recovery of protozoan cysts, coccidian oocysts, microsporidial spores, and helminth eggs and larvae (Box 49-2). Both flotation and sedimentation methods are available; the most common procedure is the formalin-ethyl acetate sedimentation method (formerly called the formalin-ether method). The concentrated specimen is examined as a wet preparation, with or without iodine, using low and high dry magnifications (xlOO, x400) as indicated for the direct wet smear examination.

The third part of the O&P examination is the permanent stained smear, which is designed to facilitate identification of intestinal protozoa (Box 49-3). Several staining methods are available; the two most common methods are the Wheatley modification of the Gomori

BOX 49-3 Permanent Stained Smear—Review

PRINCIPLE:

To provide contrasting colors for the background debris and parasites present; designed to allow examination and recognition of detailed organism morphology under oil immersion examination (100x objective for a total magnification of x1000). Designed primarily to allow recovery and identification of the intestinal protozoa SPECIMEN:

Any stool specimen that is fresh or preserved in PVA (mercuiy- or non-mercury-based), SAF, MIF, or the newer slngle-vial-system fixatives REAGENTS:

Trichrome, iron-hematoxylin, modified iron-hematoxylin, polychrome IV, chlorazol black E stains, and their associated solutions; dehydrating solutions (alcohols and xylenes or xylene substitutes); mounting fluid optional. Absolute alcohol (100% ethanol) is preferred, rather than using the 95%/5% alcohol option for absolute alcohol EXAMINATION:

Oil immersion examination (x1000) of at least 300 fields; additional fields may be required if suspect organisms have been seen in the wet preparations from the concentrated specimen. RESULTS AND LABORATORY REPORTS:

The majority of the suspect protozoa and/or human cells could be confirmed by the permanent stained smear. These reports should be categorized as "final" and would be reported as such (the direct wet smear and the concentration examination would provide "preliminary" results)

PROCEDURE NOTES AND LIMITATIONS:

The most commonly used stains include trichrome and iron-hematoxylin. Unfortunately, helminth eggs and larvae will generally take up too much stain and may not be identified from the permanent stained smear. Also, coccidian oocysts and microsporidian spores require other staining methods for identification. Permanent stained smears are normally examined under oil immersion examination (x1000), and low or high dry power is not recommended. The slide may be screened using the newer 50x or 60x oil immersion objectives but should not be reported until examination has been completed using the 100x oil immersion lens. Confirmation of the intestinal protozoa (both trophozoites and cysts) is the primary purpose of this technique IMPORTANT REMINDER:

When using non-mercury fixatives or one of the single-vial options (usually zinc-based, proprietary formula), the iodine-alcohol step can be eliminated. After drying, the slides can be placed directly into the stain (trichrome or hematoxylin). However, when material arrives for proficiency testing (e.g., College of American Pathologists, American Association of Bioanalysts), these fecal specimens may have been preserved using mercury-based fixatives. Therefore, you must include the iodine-alcohol step and subsequent rinse steps for mercury removal in the routine staining protocol. Some laboratories have decided to leave the staining protocol as is; the use of the iodine-alcohol step will not harm smears preserved using non-meroury-based fixatives

Charcot Leyden Crystals Stool Test
Figure 49-1 Charcot-Leyden crystals; stool material stained with Wheatley's trichrome stain.
Recall Leukocytes Stool
Figure 49-2 Polymorphonuclear leukocytes; stool material stained with Wheatley's trichrome stain.

important procedure performed for the identification of intestinal protozoan infections; the permanent stained smears are examined using oil immersion objective?' (x600 for screening, xlOOO for final review of 300 or more oil immersion fields).

Modified acid-fast stains are recommended for the. intestinal coccidia (Box 49-4), and modified trichron stains are recommended for the intestinal microspor (Box 49-5). These stains are specifically designed allow identification of the coccidian oocysts and microf sporidian spores, respectively.5,6

tissue trichrome and the iron-hematoxylin stains (Figures 49-1 to 49-3). This part of the O&P examination is critical for the confirmation of suspicious objects seen in the wet examination and identification of protozoa that might not have been seen in the wet preparation. The permanent stained smear is the most

Recovery of Tapeworm Scolex. Because the cation used for treatment of tapeworms is usually ve: effective, the procedure for recovery of the tapeworrffij scolex is rarely requested and is no longer clinically relevant. However, stool specimens may have to be examined for the presence of scolices and gravid ] glottids of cestodes for proper species identification, lb»

Blastocystis Hominis
Figure 49-3 Blastocystis hominis central body forms (larger objects) and yeast cells (smaller, more homogeneous objects); st 1 material stained with Wheatley's trichrome stain.

procedure requires mixing a small amount of feces with water and straining the mixture through a series of wire screens (graduated from coarse to fine mesh) to look for «polices and proglottids. Appearance of scolices after therapy is an indication of successful treatment. If the ■»colex has not been passed, it may still be attached to the mucosa; the parasite is capable of producing more segments from the neck region of the scolex, and the infection continues. If this occurs, then the patient can 1 be retreated when proglottids begin to reappear in the :,t00l.fi

Examination for Pinworm. A roundworm parasite that has worldwide distribution and is commonly found i1 children is Enterobius vermicularis, known as pinworm i seatworm. The adult female worm migrates out of the >nus, usually at night, and deposits her eggs on the perianal area. The adult female (8 to 13 mm long) may Occasionally be found on the surface of a stool specimen or on the perianal skin. Because the eggs are usually deposited around the anus, they are not commonly found in feces and must be detected by other diagnostic techniques. Diagnosis of pinworm infection is usually based on the recovery of typical eggs, which are described as thick-shelled, football-shaped eggs with one slightly flattened side. Each egg often contains a fully developed embryo and will be infective within a few hours after being deposited (Figure 49-4).6,7J7

Sigmoidoscopy Material. Material obtained from sigmoidoscopy can be helpful in the diagnosis of amebiasis that has not been detected by routine fecal examinations; however, a series of at least three routine stool examinations for parasites should be performed for each patient before sigmoidoscopy examination is done. The specimen should be processed immediately. Three

BOX 49-4 Modified Acid-Fast Permanent Stained Smear—Review

PRINCIPLE:

To provide contrasting colors for the background debris and parasites present; designed to allow examination and recognition of acid-fast characteristic of the organisms under high dry examination (40x objective for a total magnification of x400). Designed primarily to allow recovery and Identification of intestinal coocidian oocysts. Internal morphology (sporozoites) will be seen in some Cryptosporidium oocysts under oil immersion (x1000 magnification), while Cyclospora oocysts contain no specific internal morphology SPECIMEN:

Any stool specimen that is fresh or preserved in formalin, SAF- or the newer single-vial-system fixatives

REAGENTS:

Kinyoun's acid-fast stain, modified Ziehl-Neelsen stain and their associated solutions; dehydrating solutions (alcohols and xylenes or xylene substitutes); mounting fluid optional; the decolorizing agents are less intense than the routine acid-alcohol used in routine acid-fast staining (this fact is what makes these procedures "modified" acid-fast procedures). The recommended decolorizer is a 1% to 3% sulfuric acid, no stronger; many laboratories are using 1 % so the Cyclospora oocysts retain more of the color EXAMINATION:

High dry examination (x400) of at least 300 fields; additional fields may be required if suspect organisms have been seen but are not clearly acid-fast

RESULTS AND LABORATORY REPORTS:

The identification of Cryptosporidium and /sospo/a oocysts should be possible; Cyclospora oocysts, which are twice the size of Cryptosporidium oocysts, should be visible but tend to be more acid-fast variable. Although microsporidla are acid-fast, their small size will make their recognition difficult Final laboratory results would depend heavily on the appearance of the quality control (X) slides and comparison with positive patient specimens PROCEDURE NOTES AND LIMITATIONS: Both the cold and hot modified acid-fast methods are excellent for the staining of coccidian oocysts. Some believe that the hot method may result in better stain penetration, but the differences are probably minimal. Procedure limitations are related to specimen handling (proper centrifugation speeds and time, use of no more than two layers of wet gauze for filtration, and complete understanding of the difficulties in recognizing microsporidial spores). There is also some controversy concerning whether the organisms lose the ability to take up acid-fast stains after long-term storage in 10% formalin. The organisms will be more difficult to find in specimens from patients who do not have the typical, watery diarrhea (more formed stool contains more artifact material)

methods of examination can be performed. All three are recommended; however, depending on the availability of trained personnel, proper fixatives, or the amount of specimen obtained, one or two procedures may be used. Many physicians who perform sigmoidoscopy procedures do not realize the importance of selecting the proper fixative for material to be examined for parasites. Even the most thorough examination will be meaningless if the specimen has been improperly prepared.6

BOX 49-5 Modified Trichrome Permanent Stained Smear—Review

PRINCIPLE:

To provide contrasting colors for the background debris and parasites present; designed to allow examination and recognition of organism morphology under oil immersion examination (100x objective for a total magnification of x1000). Designed primarily to allow recovery and identification of intestinal microsporidial spores. Internal morphology (horizontal or diagonal "stripes" may be seen in some spores under oil immersion) SPECIMEN:

Any stool specimen that is fresh or preserved in formalin, SAF- or the newer single-vial-system fixatives

REAGENTS:

Modified trichrame stain (using high-dye-content chromotrope 2R) and associated solutions; dehydrating solutions (alcohols, xylenes, or xylene substitutes); mounting fluid optional EXAMINATION:

Oil Immersion examination (x1000) of at least 300 fields; additional fields may be required if suspect organisms have been seen but are not clearly identified RESULTS AND LABORATORY REPORTS:

The identification of microsporidial spores may be possible; however, their small size will make recognition difficult Final laboratory results would depend heavily on the appearance of the QC slides and comparison with positive patient specimens PROCEDURE NOTES AND LIMITATIONS:

Because of the difficulty in getting dye to penetrate the spore wall, this staining approach can be helpful. Procedure limitations are related to specimen handling (proper centrifugation speeds and time, use of no more than two layers of wet gauze for filtration, and complete understanding of the difficulties in recognizing microsporidial spores because of their small size [1 to 3.0 nmD IMPORTANT QUESTIONS FOR COMMERCIAL SUPPLIERS: Make sure to ask about specific fixatives and whether the fecal material can be stained with the modified trichrame stains and modified acid-fast stains. Also, ask whether the fixatives prevent the use of any of the newer fecal immunoassay methods now available for several of the intestinal amebae, flagellates, coccidia, and microsporidia

Duodenal Drainage. In infections with G. lamblia or Strongyloides stercoralis, routine stool examinations may not reveal the organisms. Duodenal drainage material can be submitted for examination, a technique that may reveal the parasites. The "falling leaf" motility often described for Giardia trophozoites is rarely seen in these preparations. The organisms may be caught in mucus strands, and the movement of the flagella on the Giardia trophozoites may be the only subtle motility seen for these flagellates. Strongyloides larvae will usually be very motile. Remember to keep the light intensity low.

The duodenal fluid may contain mucus, and the organisms tend to be found within the mucus. Therefore, centrifugation of the specimen is important, and the sedimented mucus should be examined. Fluorescent antibody or various immunoassay detection kits (Cryptosporidium or Giardia) can also be used with fresh or for-malinized material.

If a presumptive diagnosis of giardiasis is obtained based on the wet preparation examination, the coverslig can be removed and the specimen can be fixed witb either Schaudinn's fluid or polyvinyl alcohol (FVA) for subsequent staining with either trichrome or iron hema* toxylin. If the amount of duodenal material submitted is very small, then permanent stains can be prepared rather than using any of the specimen for a wet smear examination. Some believe that this approach provides a more permanent record, and the potential problem^ with unstained organisms, very minimal motility, and a lower-power examination can be avoided by using oil immersion examination of the stained specimen at xlOOO magnification.6

Duodenal Capsule Technique (Entero-Test). The duodenal capsule technique is a simple and convenient-method of sampling duodenal contents that eliminates the need for intestinal intubation. The technique involves using a length of nylon cord coiled inside a gelatin capsule. The cord protrudes through one end the capsule and is taped to the side of the patient's face. The capsule is then swallowed. The gelatin dissolves in the stomach, and the weighted cord is carried by peristalsis into the duodenum. The cord is attached toj, the weight by a slipping mechanism; the weight is released and passes out in the stool when the cord is retrieved after 4 hours. The mucus collected on the cord is then examined for the presence of parasites, including Strongyloides stercoralis, Giardia lamblia, Cryptosporidium spp., microsporidia, and the eggs of Clonorchis sinensis.6

Urogenital Tract Specimens

The identification of Trichomonas vaginalis is usually based on the examination of wet preparations of vaginafi and urethral discharges and prostatic secretions or urine sediment. Multiple specimens may need to be examined! to detect the organisms. These specimens are diluted with a drop of saline and examined under low powef (xlOO) and reduced illumination for the presence of,1 actively motile organisms; as the jerky motility begins tft diminish, it may be possible to observe the undulating membrane, particularly under high dry power (x400) (Figure 49-5). Stained smears are usually not necessary, for the identification of this organism. Many times we number of false-positive and false-negative results, reported based on stained smears strongly suggests thl value of confirmation by observation of motile organisms] from the direct mount, from appropriate culture media] or more sensitive direct antigen detection methods sue as the OSOM Trichomonas Rapid Test (Figure 49-6).6,25

Sputum

Although not one of the more common specimen^ expectorated sputum may be submitted for examination

Phlegm Smear Microscopic Pics
  1. Use a piece of clear (not frosted) cellophane tape approximately 4 inches (10 cm) long.
  2. Hold the tape between thumbs and forefingers with sticky side facing outward.
  3. Use a piece of clear (not frosted) cellophane tape approximately 4 inches (10 cm) long.
  4. Before the patient has arisen from bed in the morning (preferably while the child is still asleep), press the sticky side of the tape against the skin across the anal opening with even, through pressure._
  5. Hold the tape between thumbs and forefingers with sticky side facing outward.
Seatworm Cellophane Tape Test
  1. Before the patient has arisen from bed in the morning (preferably while the child is still asleep), press the sticky side of the tape against the skin across the anal opening with even, through pressure._
  2. Gently place the sticky side of the tape down against the surface of a dear glass slide. Label the slide with the patients's name
  3. Gently place the sticky side of the tape down against the surface of a dear glass slide. Label the slide with the patients's name
Cellophane Tape Method Fungi

for parasites. Organisms in sputum that may be detected and may cause pneumonia, pneumonitis, or Loeffler's syndrome include die migrating larval stages of Ascaris himbricoides,, Strongyloides stercorals, and hookworm; the eggs of Paragonimus westermani; Echinococcus granulosus hooklets; and the protozoa Entamoeba histolytica, Entamoeba gingivalis. Trichomonas tenax, Cryptosporidium spp., and possibly the microsporidia. Some of the smaller organisms must be differentiated from fungi such as Candida spp. and Histoplasma capsulatum. In a Parago-

Figure 49-4 Method for collection of a cellophane (Scotch) tape preparation for pinworm diagnosis. This method dispenses with the tongue depressor, requiring only tape and a glass microscope slide. The tape must be pressed deep into the anal crack.

ntmus infection, the sputum may be viscous and tinged with brownish flecks, which are clusters of eggs ("iron filings"), and may be streaked with blood.

Induced sputa are collected after patients have used appropriate cleansing procedures to reduce oral contamination. The induction protocol is critical for the success of the procedure, and well-trained individuals are necessary to recover the organisms.

Aspirates

The examination of aspirated material for the diagnosis of parasitic infections may be extremely valuable, particularly when routine testing methods have failed to demonstrate the organisms.6 These specimens should be transported to the laboratory immediately after collection. Aspirates include liquid specimens collected from a variety of sites where organisms might be found. Aspirates most commonly processed in the parasitology laboratory include fine-needle aspirates and duodenal aspirates. Fluid specimens collected by bronchoscopy include bronchoalveolar lavages and bronchial washings.

Fine-needle aspirates may be submitted for slide preparation and/or culture. Aspirates of cysts and abscesses for amebae may require concentration by centri-fugation, digestion, microscopic examination for motile

Microscopic Trichomonas Picture
Figure 49-6 A, Rapid identification kit for Trichomonas vaginalis. B, Culture system for T. vaginalis. (A Courtesy Genzyme Diagnostics, Cambridge, Mass.)
Hydatid Sand Microscope

Figure 49-7 Echinococcus granulosus, hydatid sand (300x). Inset, T\vo individual hooklets (lOOOx).

organisms in direct preparations, and cultures and microscopic evaluation of stained preparations. Aspiration of cyst material, usually liver or lung, for die diagnosis of hydadd disease is usually performed when open surgical techniques are used for cyst removal. The aspirated fluid is submitted to the laboratory and examined for the presence of hydatid sand (scolices) or hooklets; the absence of this material does not rule out the possibility of hydatid disease, in that some cysts are sterile (Figure 49-7).

Bone marrow aspirates for Leishmania amastigotes, Trypanosoma cruzi amastigotes, or Plasmodium spp. re-

Naegleria Fowleri
Figure 49-8 Naegleria fowleri in brain tissue. Hematoxylin and eosin stain. Note the large karyosome.

quire Giemsa staining. Examination of these specimens may confirm an infection that has been missed by examination of routine blood films.

Cases of primary amebic meningoencephalitis ar rare, but the examination of spinal fluid may reveal the | causative agent, Naegleria fowleri, one of the free-In amebae (Figure 49-8).

Biopsy Specimens

Biopsy specimens are recommended for the diagnose tissue parasites.6,17 The following procedures ma] used for this purpose in addition to standard histoid gic preparations: impression smears and teased anj "squash" preparations of biopsy tissue from sk muscle, cornea, intestine, liver, lung, and brain. TtsiuflJ to be examined by permanent sections or electrp microscopy should be fixed as specified by the la| ratories that will process the tissue. In certain casefrj biopsy may be the only means of confirming a suspected garasitic problem. Specimens that are going to be psamined as fresh material rather than as tissue sections jbould be kept moist in saline and submitted to the laboratory immediately.

Detection of parasites in tissue depends in part on gpedmen collection and having sufficient material to perform the recommended diagnostic procedures. Bio, y specimens are usually small and may not be representative of the diseased tissue. Multiple tissue samples often improve diagnostic results. To optimize the yield fro* 1 any tissue specimen, examine all areas and use as many procedures as possible. Tissues are obtained from jjfcvasive procedures, many of which are very expensive and lengthy; consequently, these specimens deserve the most comprehensive procedures possible. A muscle biopsy can be obtained for the diagnosis of infection with frkhinella spp.; the specimen can be processed as a ■routine histology slide or can be examined as a "squash" preparation (Figure 49-9); the life cycle of this human tissue nematode can be seen in Figure 49-10.

Tissue submitted in a sterile container on a sterile sponge dampened with saline may be used for cultures of protozoa after mounts for direct examination or jj&ipression smears for staining have been prepared. If cultures for parasites will be made, use sterile slides for smear and mount preparation. Examination of tissue impression smears is detailed in Table 49-9.

Figure 49-9 Trichinella spp. larvae encysted in muscle.

Blood

Depending on the life cycle, a number of parasites may be recovered in a blood specimen, either whole blood, buffy coat preparations, or various types of concentrations. Although some organisms may be motile in fresh, whole blood, normally species identification is accomplished from the examination of permanent stained blood films, both thick and thin films* Blood films can be prepared from fresh, whole blood collected with no anticoagulants, anticoagulated blood, or sediment from the various concentration procedures. The recommended stain of choice is Giemsa stain; however, the parasites can also be seen on blood films stained with Wright's stain. Delafield's hematoxylin stain is often used to stain the microfilarial sheath; in some cases, Giemsa stain does not provide sufficient stain quality to allow differentiation of the microfilariae.

Thin Blood Rims. In any examination of thin blood films for parasitic organisms, the initial screen should be carried out with the low-power objective (1 Ox) of a microscope. Microfilariae may be missed if the entire thin film is not examined. Microfilariae are rarely present in large numbers, and frequently only a few organisms occur in each thin film preparation. Microfilariae are commonly found at the edges of the thin film or at the feathered end of the film because they are carried to these sites when the blood is spread. The feathered end of the film where the red blood cells (RBCs) are drawn out into one single, distinctive layer of cells should be examined for the presence of malaria parasites and trypanosomes. In these areas, the morphology and size of the infected RBCs are most clearly seen (Box 49-6).

Depending on the training and experience of the microscopist, examination of the thin film usually takes 15 to 20 minutes (a300 oil immersion fields) for the thin film at a magnification of xlOOO. Although some people use a 50x or 60x oil immersion objective to screen stained blood films, there is some concern that small parasites such as plasmodia, Babesia spp., or L. donovani

Humans

Ingestion of undercooked pork containing encysted larvae

Larvae carried via woodstream to muscles (penetrate and encyst)

Bacterial Vaginosis Facts

Bacterial Vaginosis Facts

This fact sheet is designed to provide you with information on Bacterial Vaginosis. Bacterial vaginosis is an abnormal vaginal condition that is characterized by vaginal discharge and results from an overgrowth of atypical bacteria in the vagina.

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Responses

  • daniel
    What is the purpose of adding xylene in frosted type of cellophane tape?
    5 years ago

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