{"id":1265,"date":"2013-07-01T14:57:17","date_gmt":"2013-07-01T14:57:17","guid":{"rendered":"http:\/\/phosdev.com\/todaysveterinarypractice\/?p=1265"},"modified":"2022-02-17T19:05:52","modified_gmt":"2022-02-17T19:05:52","slug":"todays-technician-diagnosis-of-internal-parasites","status":"publish","type":"post","link":"https:\/\/navc.sitepreview.app\/todaysveterinarypractice.com\/parasitology\/todays-technician-diagnosis-of-internal-parasites\/","title":{"rendered":"Diagnosis of Internal Parasites"},"content":{"rendered":"

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Oreta M. Samples, RVT, MPH, DHSc<\/em><\/p>\n

The diagnosis of internal parasites in companion animals continues to evolve. Efficient methods that allow clinicians to diagnose infections more quickly and implement treatment earlier have helped pets live longer, healthier lives. Because some internal parasites spread zoonotic disease, such advances also help protect owners.<\/p>\n

PARASITE CLASSIFICATION & IDENTIFICATION<\/span><\/h2>\n

There are 2 major categories of internal parasites that affect both dogs and cats:<\/p>\n

    \n
  1. Protozoans (eg, Giardia, Toxoplasma, Tritrichomonas<\/em>)<\/li>\n
  2. Helminths (eg, roundworms, hookworms, tapeworms).<\/li>\n<\/ol>\n

    Although most parasites are associated with a particular system of the body, due to life cycle differences, evidence of their presence may be seen in a variety of places (Table 1<\/span><\/strong>). For this reason, more than 1 method of testing may be needed to confirm parasitic presence. In addition, attention to detail with regard to health history, aseptic collection, and processing of relevant samples as well as clinical experience in the laboratory help determine diagnostic answers.<\/p>\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n
    TABLE 1. Diagnostic Testing Organized by Body System & Target Parasites<\/td>\n<\/tr>\n
    Diagnostic Test<\/td>\nParasite Classification<\/td>\n<\/tr>\n
    GASTROINTESTINAL ANALYSIS<\/td>\n<\/tr>\n
    Passive fecal flotation
    \nCentrifugal flotation<\/td>\n    		Helminth eggs
    \nProtozoan cysts<\/td>\n<\/tr>\n
    Vomit flotation<\/td>\nNematode ova (helminth)<\/td>\n<\/tr>\n
    Fecal sedimentation<\/td>\nHelminth ova (especially trematode ova)<\/td>\n<\/tr>\n
    Baermann technique<\/td>\nLungworm larvae in feces<\/td>\n<\/tr>\n
    Fecal culture<\/td>\nDifficult-to-identify eggs and cysts
    \nNematode larvae
    \nCoccidial oocysts<\/td>\n<\/tr>\n
    Stained fecal smear<\/td>\nProtozoan oocysts and trophozoites<\/td>\n<\/tr>\n
    BLOOD ANALYSIS<\/td>\n<\/tr>\n
    Direct blood examination<\/td>\nHeartworm microfilaria<\/td>\n<\/tr>\n
    Modified Knott\u2019s test<\/td>\n<\/tr>\n
    Buffy coat method<\/td>\n<\/tr>\n
    ELISA testing<\/td>\nHeartworm antigens and antibodies<\/td>\n<\/tr>\n
    Stained blood smear<\/td>\nBlood protozoa<\/td>\n<\/tr>\n
    URINE ANALYSIS<\/td>\n<\/tr>\n
    Urine sedimentation<\/td>\nHelminth ova<\/td>\n<\/tr>\n<\/tbody>\n<\/table>\n

    GASTROINTESTINAL ANALYSIS<\/span><\/h2>\n

    The gastrointestinal (GI) tract plays host to a variety of helminths and protozoan parasites. Because these parasites may mature and reproduce in different areas of the GI system, there is no single best method for ova, larvae, or adult identification.<\/p>\n

    Likewise, the method and choice of flotation solution that may be used to identify parasitic ova, oocysts, or larvae will differ based on specific gravity (SG) of the organism compared to that of the chosen flotation solution. If protozoan infection of the GI tract is suspected, different testing methods may be used to identify and differentiate between oocysts and trophozoites.<\/p>\n

    The choice of solution as well as method of analysis contributes to the success or failure of accurate analysis.\u00a0Table 2<\/span><\/strong>\u00a0lists fecal flotation solutions and the parasites identified through their use.1<\/sup><\/p>\n\n\n\n\n\n\n\n\n\n\n
    TABLE 2. Quick Guide to Solutions & Target Parasites1<\/sup><\/td>\n<\/tr>\n
    SOLUTION<\/td>\nIDENTIFIABLE PARASITES<\/td>\n<\/tr>\n
    Sodium chloride*<\/span>\u2020
    \n<\/span><\/td>\n    		Common helminths
    \nProtozoan ova and cysts<\/td>\n<\/tr>\n
    Sheather\u2019s*<\/td>\nCommon helminths
    \nProtozoan ova and cysts (particularly Cryptosporidium oocysts)<\/td>\n<\/tr>\n
    Sodium nitrate*<\/span>\u2020
    \n<\/span><\/td>\n    		Common helminths
    \nProtozoan ova and cysts<\/td>\n<\/tr>\n
    Zinc sulfate*<\/td>\nCommon helminths (particularly Giardia)
    \nProtozoan ova and cysts (particularly lungworm larvae)<\/td>\n<\/tr>\n
    Magnesium sulfate*<\/span>\u2020
    \n<\/span><\/td>\n    		Common helminths
    \nProtozoan ova and cysts<\/td>\n<\/tr>\n
    *Noneffective if flukes are suspected; use of fecal sedimentation is advised
    \n\u2020Distorts Giardia rapidly, subsequently impeding diagnosis
    \n<\/span><\/td>\n<\/tr>\n<\/tbody>\n<\/table>\n
    \n

    PARASITOLOGY PRIMER<\/h2>\n

    Protozoa Life Stages<\/h3>\n

    Cyst<\/strong>:\u00a0<\/span>Infectious form of many protozoan parasites during which they are encapsulated inside a protective wall; usually found in the feces<\/p>\n

    Oocyst<\/strong>:<\/span>\u00a0Encysted, highly resistant zygotic stage of some sporozoan parasites that may remain infective for extended periods of time<\/p>\n

    Trophozoite<\/strong>:<\/span>\u00a0Active, motile feeding stage of the flagellate protozoa as well as the postsporozoite state that is seen in some apicomplexan parasites (Figure 1<\/span><\/strong>)<\/p>\n

    Adult:<\/strong>\u00a0Mature form of protozoan life capable of sexual or asexual reproduction<\/p>\n

    Helminth Life Stages<\/span><\/strong><\/h3>\n

    Ova<\/strong>:<\/span>\u00a0Scientific term for cells commonly referred to as \u201ceggs\u201d; singular is \u201covum\u201d<\/p>\n

    Larva<\/strong>:<\/span>\u00a0Independent, immature (ie, juvenile) worm that must undergo maturation in form and size to achieve adulthood<\/p>\n

    Adult<\/strong>:\u00a0<\/span>Mature helminth life form capable of reproduction<\/p>\n

    Helminth Types<\/span><\/strong><\/h3>\n

    Trematode<\/strong>:\u00a0<\/span>Of the class Trematoda, a parasite unique in its flat-bodied appearance (ie, liver fluke) and ability to parasitize certain organs, such as the liver (Figure 2<\/span><\/strong>)<\/p>\n

    Cestode<\/strong>:\u00a0<\/span>Of the class Cestoda, a parasite with a ribbon-like flattened body and specialized \u201chold-fast\u201d organ of attachment at one end; commonly referred to as tapeworms (Figure 3<\/span><\/strong>)<\/p>\n

    Nematode<\/strong>:<\/span>\u00a0Of the class Nematoda, a parasite with an unsegmented and tubular, elongated body; commonly referred to as roundworms (Figure 4<\/span><\/strong>).<\/p>\n

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    Figure 1. Trophozoites in liver tissue section<\/p><\/div>\n

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    Figure 2. Ova of Fasciola hepatica<\/em>, also known as common liver fluke (trematode)<\/p><\/div>\n

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    Figure 3. Ova of Taenia<\/em> species, also known as tapeworms (cestode)<\/p><\/div>\n

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    Figure 4. Ova of Toxocara cati<\/em>, also known as feline roundworm (nematode)<\/p><\/div>\n<\/div>\n

    Fecal Flotation<\/strong><\/h3>\n

    Fecal flotation remains the most common method to detect helminth eggs and protozoan cysts.<\/p>\n

    Flotation Solutions<\/span><\/strong><\/h3>\n

    Based on the SG differences of the various parts of a fecal sample (feces, ova, cysts, debris), the recovered eggs are lighter (ie, lower SG) than the flotation solution and will float to the surface. The heavier fecal matter (ie, higher SG) sinks rapidly.<\/p>\n

    Therefore, the flotation solution must have a higher SG than the parasite eggs or cysts. It is important to note that while water may often be a part of a fecal flotation solution, pure tap water alone cannot be used because the eggs and oocysts are heavier, sinking in the tap water and remaining unidentified.<\/p>\n

    There are several commonly used fecal flotation solutions that clinicians can quickly make for clinical use (Table 3<\/span><\/strong>).1<\/sup>\u00a0Choose solutions based on health history and expected findings\u00a0(Table 2<\/strong><\/span>).1<\/sup><\/p>\n\n\n\n\n\n\n\n\n\n\n
    TABLE 3. Fecal Flotation Solutions1,2<\/sup><\/td>\n<\/tr>\n
    SOLUTION<\/td>\nSPECIFIC GRAVITY<\/td>\nFORMULATION<\/td>\n<\/tr>\n
    Sodium chloride<\/span>*
    \n
    \n<\/span><\/td>\n    		1.2<\/td>\nAdd table salt to boiling water until the salt no longer dissolves<\/td>\n<\/tr>\n
    Sheather\u2019s<\/span>\u2020
    \n<\/span><\/td>\n    		1.2\u20131.25<\/td>\n454 g (1 lb) sugar + 355 mL water
    \n+ 6 mL 40% formaldehyde (or 1 g crystalline phenol)<\/td>\n<\/tr>\n
    Sodium nitrate<\/td>\n1.2\u20131.33<\/td>\n315 g sodium nitrate: 1 L water<\/td>\n<\/tr>\n
    Zinc sulfate<\/td>\n1.18<\/td>\n386 g zinc sulfate: 1 L water<\/td>\n<\/tr>\n
    Magnesium sulfate<\/span>
    \n<\/span><\/td>\n    		1.32<\/td>\n350 g Epson salts: 1 L water<\/td>\n<\/tr>\n
    *Sodium chloride solution, although widely used due to its low cost, is highly corrosive and will distort eggs
    \n\u2020Due to high sugar content, Sheather\u2019s solution is sticky and will attract insects if not kept sealed and the surrounding area kept clean.<\/td>\n<\/tr>\n<\/tbody>\n<\/table>\n

    Passive Fecal Flotation<\/strong><\/p>\n

    Passive fecal flotation is conducted by mixing fecal matter and flotation solution in a small vial to form a slurry; the vial is then filled until a positive meniscus forms (Figure 5<\/span>). A cover slip is placed over the meniscus and left undisturbed for 10 minutes, which allows the eggs\/cysts to float to the top (Figure 6<\/span>). The cover slip is then placed on a microscope slide and scanned at low power for signs of ova or larvae.2<\/sup><\/p>\n

    Centrifugal Flotation\u00a0<\/span><\/strong><\/h3>\n

    Centrifugal flotation \u201cspins down\u201d fecal debris, allowing the eggs\/cysts to float to the solution\u2019s surface. Slide preparation is based on the type of centrifuge used (fixed head versus swing head).<\/p>\n

    There are several centrifugal fecal flotation devices currently available that include a sieve to strain the mixture, which provides a cleaner sample for analysis; these devices include the Fecatector (butlerschein.com), Fecalyzer (vetoquinolusa.com), and Ovassay Plus (zoetis.com).<\/p>\n

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    Figure 5. Positive meniscus<\/p><\/div>\n

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    Figure 6. Cover slips placed over menisci for fecal flotation test<\/p><\/div>\n

    \n

    Passive versus Centrifugal Flotation<\/span><\/strong><\/h3>\n

    Numerous published studies have established that centrifugation is the preferred technique for parasite identification,3-6 and is, therefore, the technique recommended by most parasitologists. The centrifugal force allows a more rapid, efficient, and thorough separation of eggs\/cysts and debris.<\/p><\/blockquote>\n

    VOMIT FLOTATION<\/h2>\n

    While not common, it is possible to identify some nematode ova by evaluating vomit using the same methodology as for fecal flotation. Likewise, vomit may also be scrutinized under a microscope to locate parasites common to the stomach.<\/p>\n

    Vomit flotation is useful when parasites, such as\u00a0Physaloptera<\/span><\/em>\u00a0species or\u00a0Ollulanus tricuspis<\/span><\/em>, are suspected in dogs and cats.2<\/sup>\u00a0Vomit flotation should also be included in the initial workup for companion animals that present with chronic vomiting.2<\/sup><\/p>\n

    FECAL SEDIMENTATION<\/span><\/h2>\n

    The fecal sedimentation test is used to detect ova that have a SG higher than commonly used flotation solutions and, therefore, do not readily float. This method may also be used for ova that will be distorted or destroyed in the presence of a super saturated salt solution.<\/p>\n

    The sedimentation test is most valuable for identifying fluke ova, which have a higher SG and are larger, denser, and heavier than other ova, and should be performed in suspected cases of infection.1<\/sup>\u00a0Also, because some nongastrointestinal inhabitants, such as\u00a0Paragonimus kellicotti<\/span><\/em>\u00a0(lung flukes), may also shed ova in sputum or other internal fluids, microscopic examination should be considered.2<\/sup><\/p>\n

    BAERMANN TECHNIQUE<\/span><\/h2>\n

    The Baermann testing method (often referred to as the \u201cBaermann technique\u201d) involves the concentration of nematode larvae from a fecal sample. Although this technique is useful for identification of most nematodal larval forms, it is the gold standard method of testing for suspected cases of lungworm infection.\u00a0Table 4<\/span>\u00a0lists the most common lungworm species seen in domestic animals.7,8<\/span><\/sup><\/p>\n\n\n\n\n\n\n\n\n\n\n\n\n\n
    TABLE 4. Common Lungworm Species Seen in Domestic Animals<\/span><\/td>\n<\/tr>\n
    Lungworm Species<\/td>\nDomestic Animal Host<\/td>\n<\/tr>\n
    Aelurostrongylus abstrusus<\/td>\nCats<\/td>\n<\/tr>\n
    Capillaria aerophila<\/td>\nCats<\/td>\n<\/tr>\n
    Dictyocaulus arnfieldi<\/td>\nDonkeys and horses<\/td>\n<\/tr>\n
    Dictyocaulus eckert<\/td>\nDeer<\/td>\n<\/tr>\n
    Dictyocaulus viviparus<\/td>\nCattle<\/td>\n<\/tr>\n
    Metastrongylus apri<\/td>\nPigs<\/td>\n<\/tr>\n
    Muellerius capillaris<\/td>\nSheep and goats<\/td>\n<\/tr>\n
    Oslerus osleri<\/td>\nDogs<\/td>\n<\/tr>\n
    Protostrongylus rufescens<\/td>\nSheep and goats<\/td>\n<\/tr>\n<\/tbody>\n<\/table>\n

    Successful larval collection relies on the fact that most nematode species lack the ability to swim against gravity. When lungworms are present in the lower respiratory tract, larvae within the airways move with sputum to the pharynx where they are swallowed and later shed in the feces. Thus, in the case of lungworms, no eggs are found in the feces, only larvae.7<\/span><\/sup>While there are several variations to the treatment of the sample in terms of staining and observation, the basic principle for the Baermann technique remains rooted in the simple apparatus required.<\/p>\n

    Step by Step: Baermann Technique<\/span><\/strong><\/p>\n

      \n
    1. Break up fecal specimen and wrap loosely in cheesecloth.<\/li>\n
    2. Place specimen in tea strainer that is suspended over funnel.<\/li>\n
    3. Clamp rubber tube closed (tube is attached to end of funnel).<\/li>\n
    4. Pour lukewarm water into the funnel, covering cheesecloth packet.<\/li>\n
    5. Allow specimen to sit undisturbed overnight.<\/li>\n
    6. The next day, slowly relax clamp to collect a drop of fluid on a microscope slide. Add 1 drop of Lugol\u2019s iodine to stain larvae and view (10\u00d7) with cover slip.<\/li>\n
    7. Alternatively: Release 1 to 2 drops of fluid into small test tube. Add 1 drop of Lugol\u2019s iodine, tap mixture, place drop on slide and, with aid of cover slip, view at 10\u00d7.<\/li>\n<\/ol>\n
      \n
      \n

      Step by Step: Fecal Sedimentation Test2<\/span><\/sup> (Figure 7)<\/span><\/h3>\n
        \n
      1. Mix 2 g of feces with water in a cup (A<\/span>); remove debris by straining mixture through gauze or cheesecloth into a centrifuge-safe tube (B<\/span> and C<\/span>).<\/li>\n
      2. Balance tube and centrifuge at 1500 rpm for 5 minutes (D<\/span>).<\/li>\n
      3. Pour off liquid without disturbing the sediment.<\/li>\n
      4. Pipette a small amount from the top layer of sediment (E<\/span>) onto a microscope slide; apply a cover slip for viewing. If sediment is too dense and visibility is obscured, dilute with a drop of water before examining.<\/li>\n<\/ol>\n

        Notes:<\/p>\n