Dermatology 101: Increasing Efficiency and Enhancing Patient Diagnostics. Continuing education. Missy Streicher, AAS, CVT, VTS (Dermatology)

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Continuing education Dermatology 101: Increasing Efficiency and Enhancing Patient Diagnostics Missy Streicher, AAS, CVT, VTS (Dermatology) This program was reviewed and approved by the AAVSB RACE

Please contact the AAVSB RACE program if you have any comments/concerns regarding this program s validity or relevancy to the veterinary profession.

1 Continuing education Dermatology 101: Increasing Efficiency and Enhancing Patient Diagnostics Missy Streicher, AAS, CVT, VTS (Dermatology) This program was reviewed and approved by the AAVSB RACE program for 1 hour of continuing education in jurisdictions which recognize AAVSB RACE approval. Please contact the AAVSB RACE program if you have any comments/concerns regarding this program s validity or relevancy to the veterinary profession. Objective: After reading this article, participants will be able to obtain an accurate history, enhance current dermatology sampling techniques, and aid the veterinarian in diagnostics. Glancing at the day s schedule, Betsy the bulldog needs her ears checked and Sunny the golden retriever has a hot spot again. In most practices, at least one patient will present daily with a skin issue. Prepping for Patient Dermatology Visits When triaging patients, a technician is usually responsible for taking a TPR (temperature, pulse, respiration) while simultaneously obtaining a history from the owner based on the presenting complaint. The ability to take

2 a good history is helpful to the veterinarian because the information generates differential diagnoses before examining the patient. With dermatologic issues being so prevalent, taking an excellent dermatologic history is very important. Based on the history and physical exam, routine diagnostics should be recommended and performed. This provides excellent patient care, yields a definitive diagnosis of the type of secondary infection, creates a more defined list of the underlying cause, is good medicine, and is financially beneficial to the hospital by generating income from in house diagnostics. The hospital most likely has everything needed to complete these diagnostics. With practice, these can be done quickly by the technician enabling the veterinarian to spend more time discussing underlying cause(s) and therapeutic options with the owner. The laboratory items needed for an initial diagnosis are: microscope, glass microscope slides, cover slips, mineral oil, cotton-tipped applicators, scalpel blades stain. These items will be used to collect what represents the minimal diagnostic database for dermatology that include skin scrapings, dermatophyte culture and skin cytology. Allergies There are multiple types of allergies including atopy, food allergy, flea allergy, contact allergy and drug allergy. Atopy means strange disease and relates to allergies to environmental pollens, such as, grasses, trees, weeds, molds as well as house dust mites. Pruritus (itch) in dogs is a common sign of allergies and veterinary dermatologists frequently manage atopic patients as environmental allergies are not considered curable, but merely controllable. The act of scratching is commonly thought of as the rear foot contacting the body. However, licking, chewing, shaking head (ears), rubbing on furniture, and dragging the belly across carpet or rough surfaces are also signs of pruritus. Allergic dogs commonly have secondary infections which can also increase the pruritus level. The distribution pattern of pruritus along with the history provides clues as to what type of allergy may be present. Combinations of allergic diseases are common, such as, atopy and flea allergy; atopy and food allergy; or the co-existence of all three problems with or without secondary complicating infections. When dogs first show signs of having an allergy, regardless of type, erythema (redness of the skin) is commonly noted when examining the pruritic areas. As the dog ages, their allergy symptoms become more severe and in the case of atopy, may start seasonally then become non-seasonal. Additionally, the dog usually suffers from more intense pruritus as a consequence of increasing the summation of allergens thus exceeding the pruritic threshold. Commonly, secondary infections enhance inflammation as another source increasing pruritus in addition to the allergy. Secondary Dermatological Conditions To assess a potential secondary skin issue, look for signs of folliculitis (inflammation of the hair follicle). Examples of lesions associated with folliculitis are macules, papules, pustules, epidermal collarettes, crusts, excoriations, and alopecia. The three differentials of folliculitis are bacterial dermatitis, demodicosis and dermatophytosis. Malassezia pachydermatis (yeast) is often found in allergic patients, but does not have to be associated with a lesion. Ears, paws and face (particularly the lower lip margin) are areas often affected with Malassezia but can be found anywhere on the patient. A suspicion of secondary infection for a patient should be investigated if a dog previously responsive to steroid administration for the control of pruritus is now partially or non-responsive to steroid therapy. Referral to a veterinary dermatologist for long-term maintenance therapy may be warranted to manage the pet s allergies to minimize severity and frequency of the allergic flare-ups and in turn controlling the infections. Collecting Samples For folliculitis lesions, samples for the minimal dermatology data base are typically collected clean to dirty starting with plucking of hairs for dermatophyte culture, next collect skin cytologies and lastly the skin scrapings. Skin scrapings are generally collected from the same affected areas as the dermatophyte culture and cytology samples. These scrapings leave a residue of blood and mineral oil after collection, which may make the other samples not as informative if collected after the scraping. Obtaining a Culture for Dermatophytosis Ideally, a sample is collected for the presence of a dermatophyte (ringworm) when folliculitis and easily epilating hair are noted on a patient. Cost may be a limiting factor for this diagnostic test with bacteria being the most likely culprit of folliculitis. This diagnostic test may be postponed until a future visit; particularly if bacteria or yeast is found on skin cytology. If tasked with collecting a sample for a culture, pluck the hairs, crust and scale from the periphery or leading margin of a lesion as Dermatophytosis begins with a central lesion and migrates outward. The dermatophyte may resolve in the area it began, or at the center of the lesion, and a false negative may result if not collected from the appropriate area. The McKenzie toothbrush technique is recommended when trying to identify an asymptomatic carrier or rechecking a patient that antifungal treatment has been instituted to assess response to therapy. 2 This is accomplished by using a sterile toothbrush and combing the animal nose to tail to collect hair and scale. If areas of folliculitis are noted, the lesional area should be sampled last to prevent the spread of potential contaminants to other parts of the body. If dermatophyte cultures are performed in the clinic, use sterile scissors to cut the tips of the toothbrush and gently apply to the dermatophyte media. If color change to dermatophyte media is noted, apply acetate tape to the colony and with the aid of lactophenol cotton blue stain to identify the type of dermatophyte present with the aid of mycology identification guides.1 Contaminant organisms may attribute for media color change making proper identification with a mi-

3 Dermatology 101, continued Allergy Type Breed Predilection Age Distribution Pattern Atopy (allergies to grasses, Trees, weed, molds, dust Mites) Terrier breeds Brachiocephalic breeds Retrievers (Golden and Labrador) German Shepherd Dogs Hounds (Basset, Beagle) 6 months to 6 years With first symptoms noted between 1-3 years in approx Paws/Legs Face Ears Axillae Inguinal Any Breed (purebred or mixed) Food Allergy Labradors Spaniels (Cocker, Springer) German Shepherd Dogs Any Breed (purebred or mixed) Less than 1 year Old age Can occur at any age Same as atopic Perianal region Pruritus of mouth after eating GI issues (multiple bowel movements per day, loose stool, gaseous) Flea Allergy Any Breed Any age but may note seasonal exacerbations with increase of flea population noted Dorso-lumbar region Tail head Caudal/medial thighs Inguinal Contact Allergy Short coated dogs Thinly haired regions Any age; repeated and prolonged contact with offending allergen or chemical. Most rare Paws (interdigital spaces) Axillae Scrotum Inguinal region/belly Chin/muzzle Figure 1. Common Signalment and Distribution Patterns croscope necessary. Alternatively, the sample may be submitted to an outside lab at the onset of sample collection or after color change is noted on the media for identification of the dermatophyte organism. The type of dermatophyte present will help reveal the source so the cause can be controlled or eliminated as there is potential zoonosis. Microsporum gypseum is found in the soil, Microsporum canis is attributed to cats, and Trichophyton mentagrophytes is found in rodents. 1 Direct Impression Smear There are two techniques to collect skin cytology samples. First is a direct impression smear. 2 (Figure 2) This is accomplished by pressing the slide against the skin. Although simple, there are a few ways to get a better diagnostic sample by altering technique. Hold the glass microscope slide in the middle to decrease the torque applied to the slide when pressing against the skin. This technique will decrease the likelihood for breaking the slide, which may result in cutting the patient or the person collecting the sample. Pinch up the area of skin that will be sampled and place the slide on the pinched up area with the contact area being toward the end of the slide. Make contact with the sample area by applying steady pressure and then swirling or dragging the slide across the region. If there is crust or scale present, a need to find out what is causing the lesion is maximally ideal. Using a side of the slide, gently lift the side of

4 Figure 2. Direct impression smear note that area of interest is held by pinching and the slide is held in the middle Figure 3. Application of acetate tape to interdigital space Figure 4. Slide with new methylene blue stain prior to application of clear acetate tape Figure 5. Acetate tape preparation after paper towel has been pressed to slide to eliminate excess stain. This slide is ready for immediate viewing with a microscope. Figure 6. Skin cytology sample after staining with Dif Quik. The blue box indicates Malassezia pachydermatis; the yellow box a pair of cocci bacteria the crust (removing the entire crust may be painful to the patient) or scraping some of the scale away is beneficial for sample collection. Pustules easily rupture by using the same sampling technique as above. If the pustule does not rupture, consider using the corner of the slide to flick the top of the pustule allowing a disruption of its integrity and repeat the same sampling technique. Using a needle to rupture the pustule may cause unnecessary pain to the patient and possibly causing the skin to bleed obscuring the sample evaluation. Do not heat fix skin cytology samples as over heat fixing may cause artifact of how cells appear. Direct impression smears can be used for most skin cytology samples, but are particularly useful for exudative areas. Once the slide has air-dried, use Dif Quik stain (Wright s stain) according to manufacturer s recommendations. Using bibulous paper to blot the sample may give a false interpretation of results because of potential removal of sample onto the paper. Tape Method The second technique for collecting skin cytology samples is by using clear acetate tape. The tape must be clear if one cannot see though the tape, it will not be possible to assess the information from the tape preparation accurately with the microscope. Hold the tape by sticking the end on the middle or index finger. Stick the adhesive side to the patient s skin and apply pressure to area with the thumb of the hand that is holding the tape. Remove tape and then reapply to same or adjacent areas several times to increase chances of finding secondary infectious organisms. When collecting a sample from the paw, apply the tape to the interdigital spaces on both the dorsal surfaces and then the same piece of acetate tape. (Figure 3) With tape preparation samples, there are two choices for staining to observe organisms. The simplest method is to draw into a syringe a small amount onto a microscope slide as demonstrated in Figure 4. After the skin cytology is collected, place the adhesive side onto the slide and gently press tape onto the slide. This helps the tape adhere to the slide while simultaneously squeezing out excess stain to improve ease of viewing on the microscope. (Figure 5) A second way is to dip the tape the tape with hemostats or a clothespin with tape to the end of the slide. Do not dip into remove the adhesive and the sample. The disadvantage to this technique is losing the tape sample in the stain container, but it can be eventually recovered. Once appropriately stained, place the tape on a slide, use the paper towel as previously described to apply to a microscope slide. The advantage of the acetate tape preparation of cytology samples is that they are ready to read and do not require waiting for them to air dry. The downside of this collection method is it picks up hair, debris and creates many planes to focus and re-focus through on examination of the slide. Remember that a sample of the person s skin that was collecting the sample is on the end of the tape in the form of fingerprints; so be mindful to read the sample in the middle of tape prep to insure that it is the patient that is being assessed and not the taker of the sample. Scanning the slide using the 10X objective allows appraisal of more likely areas to note organisms if neutrophils and proteinaceous debris are noted before proceeding to oil immersion (100X objective). Once an area has been selected, immersion oil will need to be applied to that point of the slide. If evaluating a tape preparation, immersion oil is applied directly on the tape. The microscope condenser should be moved all the way up or open to evaluate for Malassezia and bacteria suspect areas is performed followed by examination on 100X, viewing HPF (high power fields) should be adequate to label the sample as NAF (no abnormal findings) or on a 0-4+ rating for pathological items that are present. To work in sync with the veterinarian, consider examining the same cytology samples with the clinician(s) several times

5 Dermatology 101, continued a year to compare what would constitute a score of 0-4+ to standardize reference levels. Ear Cytology Allergic otitis with the complication of secondary infection is common with atopic and food allergic animals which may be the only issue with that patient. Recurrent allergic otitis can be a source of discomfort for the patient and should be considered while collecting samples for ear cytology. Using a cottontipped applicator swab, collect material from both ears (even if only one ear is cited by the owner as being affected in the presenting complaint). Use a separate swab for each ear. To obtain the sample, gently swab a cottontipped applicator in each ear canal being careful not to lose sight of the tip of the applicator. Even though the otitis may be unilateral and the contralateral cytology sample does not reveal any infection, providing baseline documentation of both ears is important to maintain an accurate medical record and may demonstrate a developing problem which has not reached the magnitude of the contralateral ear. Additionally, a different type of infection than what was found in the affected ear may be present so treating both ears the same without appropriate documentation based on cytology may be contraindicated. After collecting each ear swab, roll samples on to a glass microscope slide appropriately labeling each as left or right. Making two identical slides with material from each ear is recommended because if rod-shaped bacteria are noted, a second slide is prepared for the necessary Gram stain without having to cause potentially more discomfort to the patient by having to re-swab to take a second sample. Due to the usually waxy nature of the ear swab material, lightly heat fixing the slide by waving a flame source under slide without creating carbon residue or char is recommended as this melts the wax and allows the stain to penetrate the exudate better. This is usually accomplished by using a lighter or a Bunsen burner. After heat fixing, stain one of the ear slides using Dif Quik as done with the direct impression smear. Once the sample has air-dried, examine the slide on 10X followed by oil immersion (100X) as described in the reading of skin cytologies above to evaluate for presence of infection, such as, bacteria (rod or cocci) or yeast using the 0 to 4+ scale. If rod-shaped Figure 7. Adult Demodex canis bacteria are noted, the second slide that was previously made should now be heat fixed and Gram stained to identify if the rods are Gram positive or negative assisting the veterinarian in therapeutic options. Skin Scrapes In addition to a patient noted to have any of the previously mentioned types of folliculitis lesions, dogs that also have comedones (blackheads), a history of steroid exposure (iatrogenic or with spontaneous hyperadrenocorticism), and dogs with short-hair coats seem to be more predisposed are suspect for Demodex mites, therefore; deep skin scrapes should be performed.1 First, place a heavy drop of mineral oil on a glass microscope slide. A new number 10 scalpel blade should be used for each patient because of potential for transfer of blood borne pathogens or diseases. Consider dulling the blade edge by running the edge across a metal surface like the edge of a stainless table. By dipping the blade in the mineral oil prior to scraping, the oil acts as a lubricant and a magnet of sorts to pick up the material of the scraping. Pinch the area of skin that is selected for the scraping because if Demodex are present, they are in the follicle and this will help bring the mites to the skin surface. After pinching, release the skin and use the non-dominant hand to spread the skin taut. Continuing to pinch the skin may act as a tourniquet allowing more scraping than necessary to see blood. This method will potentially cause more discomfort to the patient and put fingers in closer proximity to the scalpel increasing chances of a blade to finger mishap. Hold the blade perpendicular to the skin or slightly away from you. Scrape using steady light pressure in direction of hair growth ideally towards you in a small area to minimize discomfort for the patient and skin damage post sample collection. Continue to scrape until a small

6 amount of blood is noted on the blade. Skin may bruise and appear to bleed, but stopping before blood is observed on the blade may result in a false negative scrape. Once blood is noted, give the area another pinch to create more capillary ooze and the use the scalpel blade to collect one last scrape in a sweeping or scooping motion to get as much oil, blood and skin debris as possible for the sample. Pick the microscope slide with oil on it up and swirl the material into the oil - rubbing the edge on the side of the slide to remove as much material from the blade. Placing a cover slip on top of the sample is advisable to enhance recognition of the borders and ensure that the entire area has been investigated. Although Demodex mites can be visualized on 4X, 10X is preferred as the mites, particularly the eggs, are more obvious and not as easily missed. The condenser should be lowered (as done with fecal samples) to allow for increased contrast. This is particularly helpful no longer are as refractive and bright light may obscure visualization. Dead mites are still considered positive scrapes when reassessing success of therapy with a demodicide. If folliculitis lesions are present in areas that may be difficult to scrape or are risky (because of proximity to the eye for example), hair may be plucked in the direction of hair growth and then placed in mineral oil. This technique may not yield as many mites as an actual deep skin scrape, but is still informative. If Demodex mites are present, note the quantity and life stage per slide obtained and also record location of the specimen acquisition. Noting the life stages (egg, larvae, nymph, adult) is important because when a dog is treated for demodicosis, it is an adulticide over several months. If juvenile stages are noted after previously only adults are present, then finding out why that occurs is important. Examples would be if the owner is giving medication as directed, or a young dog may grow out of his therapeutic dose, or other cause such as systemic disease. (Figure 7) If Sarcoptic mange (scabies) is suspected, superficial skin scrapes are indicated. This endeavor is messy. Apply a generous amount of mineral oil to the suspect areas on the dog (margin of ear pinnae, hocks, elbows and lowest part of the ventrum) as these mites can move fast and the oil application can slow them down. Using a scalpel blade, remove the mineral oil that was placed on the skin by using long, broad strokes in the direction of hair growth and apply to a glass microscope slide. If crusts are present, scrape down to the normal skin which will not be normal due to inflammation and may bleed because of the crust removal even though causing the skin to bleed is not the objective of superficial skin scraping. Placing a cover slip on the sample, examine by scanning using the 10X objective with the condenser lowered as with the deep skin scrapes. Several slides may be required as Sarcoptes are notoriously difficult to find, but one Sarcoptes mite is considered diagnostic. Often, scabies is treated on index of suspicion (scabies incognito), but identifying them is gratifying and fulfills the diagnosis. In conclusion, technicians can collect and evaluate samples needed for the minimal database recommended for patients with dermatologic issues efficiently and effectively with practice. With skin and ear issues being seen routinely on receiving days, a technician may play a great role by evaluating the diagnostics performed thereby providing the veterinarian with a definitive diagnosis of infection type. Identifying the presence of mites allows for a more informed differential diagnosis for the underlying cause and potentially generates additional income for the hospital. References 1. Miller WH, Griffin CE, Campbell KL. Small Animal Dermatology, 7th ed. St. Louis, MO. Elsevier. Muller & Kirk Hnilica, KA. Small Animal Dermatology A Color Atlas and Therapeutic Guide, 3rd ed. St. Louis, MO. Elsevier About the Author: Missy Streicher, AAS, CVT, VTS (Dermatology) Missy has been a Certified Veterinary Technician since 1993 and worked in a small animal hospital until 2000, when she began working for a private dermatology practice. In 2006, Missy moved to the Auburn University Small Animal Teaching Hospital and continued specializing in Dermatology. She sat on the organizing committee for the Academy of Dermatology Veterinary Technicians, which was accepted as a VTS by NAVTA in July of 2015.

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