Diagnostic techniques and sample collection

Clinical Techniques in

Small Animal Practice November 2001

Vol 16, No 4

Diagnostic Techniques and Sample Collection Cathy F. Curtis, B Vet Med, DVD, MRCVS

With the advancing knowledge of medicine, diagnostic techniques and laboratory equipment, practicing veterinary surgeons are increasingly capable of processing many of their patients' blood, urine, and tissue samples on-site. No case is better suited to this scenario than the dermatology case, which enjoys a unique privilege in the clinic because the clinician is frequently able to harvest, process, and analyze pathologic samples from affected animals during or shortly after the consultation, eliminating the need to wait for results from external laboratories and consequently reducing the time to recovery. Good sampling techniques are crucial for the acquisition of optimal data however, and this article highlights the key points of these procedures and recommends when they are best employed. Copyright © 2001 by W.B. Saunders Company

Coat Brushings he simplest and probably the most frequently employed

T dermatologic diagnostic test is a coat brushing in which

hairs, scale, and debris are dislodged from the affected animal. Medium- to large-breed dogs should be made to stand or sit on a large piece of absorbent white paper while a brush or the hand is used to vigorously "comb" through the coat. Cats and small dogs can be held under one arm above the paper. By tipping the corners of the paper, the shed material can be accumulated in the center, and loose hairs can be removed for microscopic examination (see later). A scalpel blade moistened with liquid paraffin can be used to scrape up some of the scale and debris if this also needs to be examined microscopically; in the majority of cases however, this test is used to gauge an animal's flea

No reprints available. Correspondence to Cathy F. Curtis, BVetMed, DVD, MRCVS, 7 Chadwell, Ware, Hertfordshire, SG12 9JX UK. Copyright © 2001 by W.B. Saunders Company 1096-2867/01/1604-0002535.00/0 doi:10.1053/svms.2001.26998

burden, so the next step would usually involve moistening the debris with a few drops of water to determine whether the darkly colored pieces represent flea feces. If they do, a dark-red streak passes from them into the moistened tissue paper as the semidigested blood dissolves (Fig 1), whereas general debris and dirt is insoluble and remains intact. It should be noted however, that a negative coat brushing does not mean that the animal is free of fleas, because some infested individuals remove all traces of flea feces during grooming.

Skin Scrapings Another very commonly employed diagnostic test is the procurement of superficial or deep skin scrapings that comprise, respectively, the epidermal outer layers or the entire epidermis and superficial dermis. Used principally for the detection of ectoparasites (although the hyphae and spores of some nonfollicular dermatophyte species, eg, Microsporum persicolor, may be observed in scale preparauons), the habitat of the suspected parasite should be borne in mind when harvesting specimens, because this affects the depth to which the skin should be scraped. • Trombiculid mites: Reside on the skin surface with mouth-

parts embedded into the dermis. They are often visible to the naked eye, but confirmation and species identification may require microscopy so they can be removed by very superficial scrapings. • Cheyletiella sp mites: These mites live on the surface of the skin in epidermal pseudo-tunnels comprised of scale produced in response to their feeding and activity. They can be removed by superficial skin scrapings. • Otodectes cynotis mites: These mites usually reside in the vertical and horizontal ear canals of their hosts, but occasionally they escape the ear and feed on periauricular, dorsal, and distal tail skin, resulting in papular lesions. Consequently, they may be detected in periauricular tape strippings (see

Clinical Techmques in Small Animal Practice, Vol 16, No 4 (November), 2001: pp 199-206

199

2":',"

.

Fig 1. Appearance of flea feces on moistened paper towel. Note the streaks of soluble, semidigested blood that emanate from the solid pieces of feces.

below) or superficial papule scrapings, but false negatives are common because the mites are so motile. Cerumen smears may be more useful; samples can be removed from the ear using a cotton swab (Q tip), rolled on to a glass slide, and covered by a drop of liquid paraffin and a cover slip before being examined microscopically. Mites and eggs can be detected using this method. • Sarcoptes/Notoedres sp mites: Adult males and females, nymphs, and larvae may be found on the surface of the skin, but gravid females, ova, and newly emerged larvae and nymphs may be in the superficial to mid-epidermal layers (not usually proximal to the stratum granulosum). The burrowing activity of the pregnant female typically produces a papulocrustous lesion, so deep skin scrapings should be taken from atraumatized, crusted papules to ensure that all layers of the epidermis have been sampled. • Demodex sp mites: A follicular mite in both cats and dogs, it results in erythema and hair loss. Alopecic skin being sampied for demodex should be gently squeezed before scraping to encourage the mites to move distally within the hair follicle, facilitating their removal during deep scrapings. The rarer and more superficially located feline species, Demodex gatoi, which resides in the stratum corneum, may be removed by superficial scrapings. Irrespective of depth, a size-10 scalpel blade that has been dipped in liquid paraffin should be used for scraping. The site to be sampled should be scraped repeatedly in the direction of hair growth, and the material that is dislodged should be suspended in a few drops of liquid paraffin on a microscope slide. When performing deep scrapings, the skin should exude a slight ooze from dermal capillaries at the end of the procedure to ensure that the entire epidermis has been removed. A cover slip should always be applied to the sample on the slide to aid microscopic examination and to prevent motile mites from escaping. Once made, the preparation should be scanned as soon as possible at low power (40×) and any areas of interest magnified at medium power (100×). Higher magnification is not usually necessary and can be counter-productive, as it may distort the image of the mites. 200

Fig 2. Acetate tape stripping from a rat with pediculosis showing multiple louse eggs ("nits") adhering to the hairs.

Tape Strippings A relatively underused but simple and useful test, particularly for the detection of surface-resident ectoparasites, is tape stripping. The sticky side of a 10-cm length of acetate tape can be repeatedly pressed over an area suspected to be harboring a surface parasite, eg, the trunk of an animal with suspected pediculosis; the scaling dorsum of a cat, dog, or rabbit with cheyletiellosis; the hair coat of a rabbit, Guinea pig, rat, or mouse with fur mites or the periauricular area of a cat or dog with otodectic acariasis. The adherent scale, debris, and hairs removed by the tape are then trapped between it and the microscope slide and with the tape doubling up as a cover slip, the preparation is ready for immediate inspection (Fig 2). Cytologic specimens can also be obtained using a stained tape-stripping technique that is again rapid and straightforward. Lesional areas can be sampled by simply pressing the adhesive side of the tape onto the skin for a few seconds; on removal, an "imprint" of the surface will be obtained. This material actually represents the most distal layers of the stratum corneum, but in addition to the cornified squames that make up the majority of the cellular population, commensal and pathologic microbial organisms may also be harvested and can be observed if the preparation is treated with a suitable cytologic modified Wright's stain (Dill Quik) or Giemsa (Fig 3). Although it is not possible to species-type microbes harvested this

Fig 3. High-power (1,000x), oil-immersion appearance of a Diff-Quik-stained tape stripping from a basset hound with Malassezia dermatitis. Note the scores of budding yeast organisms. CATHY F. CURTIS

O

\

\

N Fig 4, Low-power (40x) trichogram in liquid paraffin depicting the hair-bulb region of anagen hairs,

Fig 5. Low-power (40x) trichogram in liquid paraffin depicting the hair-bulb region of telogen hairs.

way, a crude estimate of population size can be made (for example, more than 2 to 3 yeast organisms per high-power field may represent a significant Malassezia overgrowth, particularly in samples taken from the axillary, inguinal, and interdigital regions), and their morphology may assist the clinician with an empirical choice of antimicrobial while awaiting culture and sensitivity results.

period, and climate on this ratio can make it difficult to interpret. With experience, the relative difficulty or ease with which hair is epilated may provide us with similar information. The bulb section must also be examined for infectious agents such as dermatophytes and Demodex sp mites. Dermatophyte elements can be found both within the hair as hyphae and surrounding the hair as arthrospores (Fig 6). Their nutritional requirement for keratin means that they are more likely to invade anagen hairs that are actively producing the protein. Infected hairs are often described as having a "fuzzy" outline when examined microscopically at low (40×) or medium (100×) power. To aid visualization of the fungi, improved contrast can be obtained by reducing the aperture of the iris diaphragm. Demodex mites are the only ectoparasite intimately associated with the proximal section of the hair, owing to their intrafollicular habitat. Dogs, cats, and small mammals that resent the more lengthy process of skin scraping may tolerate hairs being plucked from lesional areas. When these samples are suspended in liquid paraffin, mites of all life-cycle stages and ova stages may be observed among the hairs (Figs 7 and 8). Occasionally, the view of the mites is obscured, but a few minutes under the light and heat of the microscope lamp may drive them away from the hairs. Thus, negative specimens should be re-examined a few minutes later. Shaft: Fungal elements may be observed within and around

Wood's Lamp Examination The Wood's lamp is an ultraviolet hght that can be used to induce a yellow-green fluorescence in some dermatophyte species. Up to 80% of Microsporum canis strains will fluoresce in this manner when illuminated for 3 to 5 minutes. More unusual fluorescent species are M. audoinii, M. distortum, and Trichophyton schoenleinii. False negatives can occur however, with a proportion of M. cauls strains and the other, commoner nonfluorescent dermatophyte species (eg, M. gypseum, T. mentagrophytes); therefore, broken hairs and scale from suspicious lesions should be examined microscopically (see below) and/or submitted to a mycologist for culture and identification. This technique is described in more detail elsewhere in this issue.

Trichoscopy The examination of a hair plucking (a trichogram) under the microscope can provide a plethora of diagnostic information. The hairs to be examined may have been dislodged during a coat brushing (see above) or forcibly removed using tweezers or hemostats, although samples obtained using the latter instruments are improved by the application of a rubber sleeve to the grasping ends to both improve contact between the hair and the forcep and to minimize hair-shaft damage during epilation. The entire length of each hair should be examined, because diagnostic information may be obtained from each of its three sections. • Bulb section: The microscopic appearance of the hair bulb from a growing or anagen hair differs from that of a hair in the resting or telogen phase of the hair cycle. Anagen hair bulbs adopt a "flared" or rounded appearance when suspended in liquid paraffin (Fig 4) and the telogen bulbs are "brush-like" (Fig 5). The ratio of anagen to telogen hairs can potentially provide some useful information on overall levels of follicular activity, but the influences of breed, age, gender, photoDIAGNOSTIC TECHNIQUES AND SAMPLE COLLECTION

Fig 6. Feline hair infected with Microsporum canis. Note the collar of spherical arthrospores surrounding the hair shaft. (Liquid paraffin; original magnification x100.)

201

Fig 7. Trichogram showing adult Demodex sp mites from a dog, suspended in liquid paraffin. (Original magnification x40.)

the hair shaft (see Bulb section above), but this section should also be examined for ectoparasites. Lice, Cheyletiella sp, and fur mites lay their eggs along the proximal to midsection of the hair shaft and attach their ova using either a cement-like substance or thread (Fig 9), although those attached by thread often become detached from the hair during sampling and may be found among the hair shafts. To differentiate between louse and mite ova, the former are always glued to the hair at the base, are larger than mite eggs and possess an operculum, through which the nymphal louse emerges (Fig 10). Adult ectoparasites may also be found in this section of the hair, particularly lice and fur mites, because they feed on hair keratin and skin debris (Fig 11). The hair shaft should also be examined for structural abnormalities such as: the defects caused by large aggregates of melanin as seen in color-dilute alopecia; mid-shaft fractures that can occur as a sequela to dermatophyte infection, self-trauma, or, rarely, as an inherited disorder such as trichorrhexis nodosa; mis-shapen and deformed hairs that can develop during periods of nutritional imbalance or may be present from birth, eg, the twisted shafts of pili torti which has been reported in a litter of kittens and keratosebaceous aggregates (known as follicular "casts"), which are a feature of certain keratinisation defects, eg, sebaceous adenitis and primary idiopathic seborrhoea. • Tips: Examination of the distal hair segments mayenable the

f ~ '

Fig 9. Cheyletiella sp mite ovum from a canine trichogram. Note the "web"-Iike thread used to attach the egg to the hair. (Original magnification x400.)

clinician to determine whether hair is being removed traumatically due to pruritic or psychogenic causes, or whether atraumatic hair loss is occurring. The normal hair tip tapers to a fine point, but traumatized hairs have fractured, split ends and can be used to betray the "secret" groomer to owners who have not observed signs of pruritus or self-trauma in their pets.

Cytology During the past decade, an increasing number of clinicians have started to harvest cytologic specimens either for in-house assessment or for submission to a specialist cytopathologist. The skin is perfectly suited to this technique because its various layers are readily accessible. N o d u l e s and Masses Solid epidermal or dermal masses should be fixed between the fingers and a 23 ° or 21 ° needle, which is attached to a 5-mL syringe, should be inserted. With the needle fixed in position, the syringe plunger should be withdrawn and then relaxed before redirecting the needle in 3 to 4 different directions within the mass and repeating the plunger movements (Fig 12).

)

j .....

.fi

~°

Y Fig 8. Triohogram from a dog with demodicosis showing 2 Demodex canis eggs nestled among the hairs. (Original magnification x 100.)

202

%

%

Fig 10. Polyplax spinulosa ovum from a rat with pediculosis. Note the thick plaque of cement substance attaching the base of the egg to the hair, and the distinctive operculum at the anterior pole. (Original magnification ×100.) CATHY F. CURTIS

\

\

Fig 11. Trichogram from a dog showing the biting louse, Trichodectes canis, attached to a canine hair. (Original magnification x40.)

With the plunger relaxed, the needle and syringe are then removed from the mass and the needle immediately detached to allow the syringe to be filled with 5 mL of air. The needle is then reattached and the air is used to expel the contents of its hub and shaft onto a microscope slide positioned beneath it. The expelled material can then be gently smeared using another glass slide, air-dried, and either submitted to a laboratory or fixed and stained with Diff-Quik for direct examination. A layer of immersion oil and a cover slip can be placed on the specimen for microscopic inspection at medium power (100 x), but areas of interest should be examined at high power (1000×), which requires a further drop of immersion oil to be placed on top of the cover slip. A detailed description of the cytologic appearance of specimens from cutaneous masses is beyond the scope

J''Lz 1

2

3

Fig 13. Pricking of an intact pustule with a sterile needle for cytologic sampling.

of this article, but the reader is referred to more detailed texts on the subject in the reading list. Pustules Whenever possible, intact pustules should be sampled, because their contents are uncontaminated by commensal cutaneous microbes. A sterile needle should be used to prick the surface of the pustule (Fig 13) and the contents can then be gently expressed and smeared across the surface of a microscope slide. Again, the sample is air-dried before being mailed to the laboratory, or fixed and stained in-house and the inspection procedure is the same as that described above for specimens from nodules and masses. The vast majority of pustules will contain a mixture of intra- and extracellular bacterial organisms (principally cocci) and a mixture of healthy and degenerate inflammatory cells (predominantly neutrophils) (Fig 14), characteristic of superficial pyoderma. Some immune-mediated diseases feature sterile pustules however, that contain no bacteria and predominantly healthy granulocytes. Sterile neutrophilic pustules can be seen in pemphigus foliaceus, an autoimmune disease characterized by autoantibody production against components of the epidermal cell desmosomes, cell desmosomes which results in detachment of the epidermal cells or acantho-

5 9 8 7

Y

Fig 12. Fine-needle aspirate biopsy technique for sampling epidermal and dermal masses. 1. Insert a 5-mL syringe with a detachable 23-gauge needle into the mass, with the plunger fully home. 2. Withdraw the plunger 2 to 3 times. 3. Relax the plunger. 4. Without removing the needle from the mass, redirect the needle within it. 5. Withdraw the plunger 2 to 3 times. 6. Allow the plunger to relax (repeat steps 4, 5, and 6 if sampling a large, solid mass). 7. Withdraw the syringe and needle. 8. Detach the needle from the syringe and fill it with air. 9. Reattach the needle and expel the air rapidly through the needle to dislodge the cells and fluid accumulated within its bore. Spray the specimen onto a clean microscope slide and smear gently before staining with a suitable cytologic stain (eg, Diff-Quik or Giemsa). DIAGNOSTIC TECHNIQUES AND SAMPLE COLLECTION

. =

Fig 14. High-power (1,OOOx), oil-immersion view of DiffQuik-stained material harvested from a pustule on the skin of a dog with superficial pyoderma. Note the predominantly neutrophilic cell population and the presence of several aggregates of intra- and extracellular coccoid organisms. 203

folds and in the case of abscesses and pyogranulomas, should be inserted in to the center of the tumor after taking care to avoid contamination with surface microbes by cleansing the surface of the skin. Tissue samples should be harvested from areas of chronic or relapsing dermal infection, and can be obtained using 4 to 6 mm skin biopsy punches or a tru-cut biopsy needle. To preserve the bacteria during transit, the tissue samples can be embedded into the transport medium at the base of a bacteriology swab container. On arrival at the laboratory, they will be macerated and inoculated onto an appropriate culture medium. Samples from cases of suspected mycobacterial infections should be submitted to a laboratory experienced in isolating and identifying such organisms, because some species are difficult to grow in vitro. Fig 15. High-power (1,000x), oil-immersion view of DiffQuik-stained material harvested from a pustule on the skin of a dog with pemphigus foliaceus. Note the larger, "rounded" acantholytic cells surrounded by predominantly healthy neutrophils in the absence of microbial organisms.

lytic cells that float among the neutrophils (Fig 15). They can also be seen in the rare disease, canine subcorneal pustular dermatosis. Eosinophilic pustules are a feature of canine sterile eosinophilic pustulosis, another rare disease. Ear Smears While awaiting the culture and sensitivity results from specimens taken from a diseased ear canal, an empirical choice of antimicrobial/anti-inflammatory can be aided by information obtained from cytologic preparations made at the time of sampiing. After harvesting material for the laboratory, a simple cotton swab can be used to collect a small amount of cerumen or pus, which should be wiped on a microscope slide. "Greasy" samples (ie, those containing a lot of cerumen) may need to be heat-fixed prior to air-drying and staining. The relative prevalence of the various cell types and microbial forms (ie, bacteria vs yeast, cocci vs rods) should be assessed.

Collection of Materials for Culture The culture and identification of bacterial, fungal, and occasionally viral organisms are important steps toward the provision of a reliable treatment regimen and prognosis for animals with infectious cutaneous diseases. Every effort should be made to yield the maximal amount of information from the samples submitted. To aid in this process, when dealing with suspected cases of unfamiliar microbes, laboratory staff should be consulted regarding lesion selection and sampling, appropriate transport media, and acceptable postage times prior to sample submission Bacteriology Samples When sampling an intact pustule, the surface of the lesion should be wiped with surgical spirit before lancing with a sterile needle. A sterile cotton swab should then be used to harvest a freshly expressed bead of pus and the swab should be mailed to the laboratory in transport medium that is capable of supporting anaerobic and aerobic bacteria, while inhibiting the growth of contaminant fungi. With intertriginous lesions, the swab should be rolled gently between the surfaces of infected skin

204

M y c o l o g y Samples

Dermatophytes Hair pluckings are the most suitable samples for dermatophyte isolation, although it should be remembered that some uncommon species (eg, Microsporum persicolor) are found only in the stratum corneum; thus, dry scrapings of skin scale should also be collected. When sampling hair, broken hairs from the center and intact hairs from the margin of an alopecic, crusting lesion should be epilated using clean tweezers and placed in a sterile container for transport to the laboratory. Alternatively, a plucking can be placed on dermatophyte test medium (DTM) for in-house culture, provided that an experienced member of staff is able to check daily for 10 days for a yellow to red color change that occurs as a result of the alkaline metabolites produced by the dermatophytes affecting the phenol red pH indicator. After 10 to 14 days, contaminant fungi can cause this color change; thus, the results are inconclusive if the plate is read after 10 days. Fungi grown on DTM or at the laboratory can be identified by the shape of their macroconidia, which can be harvested by gently pressing the sticky side of a small square of acetate tape against the surface of the colony, a procedure known as Roth's flag technique. The tape is then pressed onto a drop of lactophenol cotton blue stain, which has been placed on a glass slide and then examined microscopically.

Yeast Skin or mucosal yeast infections with organisms such as

Malassezia or Candida sp can be sampled using a sterile, moistened swab, which is then submitted to the laboratory in appropriate transport medium. An alternative method involves the use of "contact plates," which are available from some European laboratories. These are small (approximately 2.5-cm diameter) dishes of aFar capable of sustaining yeast growth and designed to be pressed against the infected skin or mucosal surface for a few seconds. The plates are then incubated for a few days in the laboratory at 32 ° to 35°C, or for a week in the clinic at 20 ° to 30°C. In the case of a Malassezia overgrowth, they will develop multiple cream-colored colonies of Malassezia pachydermatis (Fig 16). Subcutaneous (eg, pythiosis, sporotrichosis, phaeohyphomycosis) or deep (eg, cryptococcosis, histoplasmosis, blastomycosis) mycoses are uncommon to rare and generally present as single or multiple, papular to nodular lesions that may ulcerate and have draining tracts. As a group, these lesions can be sampled by swabbing the draining fluid or by skin biopsy, CATHY F. CURTIS

Fig 16. Modified Dixon's agar contact plates supporting almost-confluent growths of Malassezia pachydermatis from a West Highland white terrier with Malassezia dermatitis.

because the material harvested can be submitted for culture. Laboratory staff should be prewarned of your suspected differential diagnoses because of the human health risks involved in handling such organisms. Virology Samples Viruses are an uncommon cause of skin disease in domestic pets and lesion samples intended for virus isolation are rarely submitted to laboratories. One exception is the suspected case of the potentially zoonotic disease, feline cowpox. To make a definitive diagnosis, fresh crusts and scabs from cats presenting with historical and clinical signs suggestive of this disease should be mailed to the laboratory in viral transport medium and the staff should be forewarned that cowpox is a differential diagnosis. The majority of the other viral diseases causing dermatologic signs in small animals (eg, feline leukemia virus, feline immunodeficiency virus, feline herpes virus, and canine distemper virus) are diagnosed by the identification of viral inclusion bodies or giant cells in skin biopsy specimens (see below).

although pathologists prefer specimens with a minimal diameter of 6 mm. With the exception of facial and pedal skin, it is often possible to harvest punch biopsy specimens under the influence of a sedative using only local anesthetic as an analgesic. Once sedated, the site to be sampled should be clipped using scissors, the direction of hair growth noted, and the area outlined with a marker pen before the subcutaneous rejection of 0.5 to 1.0 mL of 1% to 2% lignocaine hydrochloride. Preparation of the site with surgical scrubs and alcohol, etc, should be avoided, because diagnostic material may be removed from the skin surface. Once the skin is anesthetised, the punch biopsy can be inserted using a combination of forward pressure and unidirectional rotational motions and should be advanced to the depth of the subcutaneous fat layer. The detached piece of skin is then gently lifted at one edge using the point of a sterile needle or fine rat-toothed forceps and any adherent tags of fat excised. Excess blood can be absorbed with a gauze swab before placing the sample on a small square of thin card, which has an arrow depicting the direction of hair growth drawn on it to aid the pathologist during sectioning. A minute later, once the sample has adhered to the card, both should be placed in a container and fixed with at least 10 times their volume of 10% phosphate-buffered formalin solution. The skin wound is closed routinely and the site can then be cleansed. Whenever possible, multiple biopsies that are representative of the spectrum of skin lesions affecting the animal should be harvested; in the case of small papules, pustules, and nodules, the lesions should be excised in toto. The pathologist should be provided with adequate details of the animal's signalment, disease history and concurrent clinical and dermatologlc signs to maximize the information he/she can supply to the veterinarian. If infectious disease is suspected (eg, dermatophytosis, leishmaniasis, my-

Skin Biopsy Techniques With the input of an experienced dermatopathologist, skin biopsy specimens can provide the clinician with crucial information, particularly when dealing with an unfamiliar clinical presentation or a case that progresses or relapses despite seemingly appropriate empirical therapy. Ironically, the pathologist is more often able to help the clinician negate certain differential diagnoses than to confirm a specifc, eg, allergic, endocrine, or ectoparasitic etiology. However, the correct classification of a disease process is the first step to a definitive diagnosis, so "rule-outs" are almost as important as "rule-ins". P u n c h Biopsy The simplest and quickest method of skin biopsy involves the use of a skin "punch", which is available in a variety of sizes, DIAGNOSTIC TECHNIQUES AND SAMPLE COLLECTION

4D-B

C

Fig 17. Punch and elliptical skin biopsy techniques for ulcerated lesions. (A) A punch biopsy specimen sectioned in this plane will contain diagnostic pathology as the ulcer margin and junctional zone between ulcerated and non-ulcerated skin will be included. (13} Specimens sectioned in this plane, however, may not contain any diagnostic features, because the junctional zone has not been sampled. (C) To avoid the problems in (B), harvest elliptical biopsies from ulcerated lesions, because the pathologist will section through the long axis of the sample and will not miss the junctional zone. 205

cobacterial infection), special stains should be requested at the time of submission. Elliptical Biopsy Particularly useful for sampling ulcerated lesions, the elliptical biopsy enables the pathologist to align the specimen along its long axis for sectioning, ensuring that the junctional zone between ulcerated and non-ulcerated skin is not lost during processing (Fig 17). Provided that the ellipse is not too large, the procedure can again be performed under sedation and local anesthetic, but larger specimens would require general anesthesia. Sample preparation and fixation is as stated above. Excisional Biopsy If it becomes necessary to remove an entire epidermal or dermal mass for health or cosmetic reasons, a routine excisional biopsy procedure can be performed and a peripheral margin of healthy tissue should be included wherever possible. The mass should be fixed as above and submitted as a whole for histopathologic examination. Large masses should be transected at 1-cm intervals to allow adequate penetration of the formalin solution.

References and Further Reading Ackerman LJ: Diagnostic tests, in Pratt P (ed): Practical Feline Dermatology (ed 2). Goleta, CA, American Veterinary Publications Inc, 1989, pp 7-37

206

Baker R, Lumsden JH: Colour Atlas of Cytology of the Dog and Cat. London, England, Mosby, 2000 Bowman DW: Arthropods, in Georgi's Parasitology for Veterinarians (6th ed). London, England, Saunders, 1995, pp 1-82 Littlewood J: Investigative and laboratory techniques, in Locke PH (ed): Manual of Small Animal Dermatology. Cheltenham, England, BSAVA Publications, 1993, pp 33-44 Moriello KA, Mason IS: Diagnostic testing, in Handbook of Small Animal Dermatology. Oxford, England, Pergamon, 1995, pp 19-44 Nesbitt GH, Ackerman LJ: Fundamentals of dermatological diagnosis, in Dermatology for the Small Animal Practitioner. Veterinary Learning Systems Co Inc, 1991, pp 13-44 Paterson S: Introduction, in Skin Diseases of the Dog. Oxford, England, Blackwell Science, 1998, pp 1-18 Paterson S: Introduction, in Skin Diseases of the Cat. Oxford, England, Blackwell Science, 2000, pp 1-20 Perman V, Alasker RD, Riis RC: Cytology of the Dog and Cat. American Animal Hospital Association, Indiana, 1979 Quinn PJ, Carter ME, Markey BK, et al: General procedures in microbiology, in Clinical Veterinary Microbiology (ed 3). London, England, Mosby, pp 9-117 Quinn PJ, Carter ME, Markey BK, et al: Dermatophytes, in Clinical Veterinary Microbiology (ed 3). London, England, Mosby, pp 381-390 Scott DW, Miller WH, Griffin CE: Diagnostic methods, in Muller and Kirk's Small Animal Dermatology (ed 6). Philadelphta, PA, Saunders, 2000, pp 71-206 Tyler RD, Cowell RL, Meinkoth JH: Cutaneous and subcutaneous lesions: Masses, cysts, ulcers and fistulous tracts, in Cowell RL, Tyler RD, Meinkoth JH (eds): Diagnostic Cytology and Haematology of the Dog and Cat (ed 2). London, England, Mosby, 1999, pp 20-51 Williamson GT, Harvey RG: Diagnostic tests and clinical pathology, in Colour Atlas of Small Animal Dermatology--A guide to dtagnosis (ed 2). London, England, Wolfe, 1994, pp 33-52

CATHY F. CURTIS