CASE REPORT

Forty-five year-old female who had presented lateral-distal right halux onycholysis for 8 months with two negative mycological tests (direct and culture). Clipping was performed (Figure 1).

Histopathological report:

Histological sections of nail plate showing parakeratosis areas, some serous lakes and hyperkeratosis of ventral area. Absence of inflammatory cells. The investigation for fungi was positive, with presence of sparse clusters of hyphae irregularly distributed, with delicate walls of variable diameters, suggestive of non-dermatophytic fungus (Figure 2).

Special staining – Resistant PAS-diastase

Do you know how to make a clipping?

It is very easy to perform it in the office. Simply cut the fragment of the distal part of the affected nail at least 5 mm long in the horizontal direction of the plate. Even though some authors think that 3 mm is enough ¹, in our experience, 5 mm or more is ideal (Figure 3). The fragment width is equally important, which should be at least 2 mm, because fixation in paraffin depends on these details. If nail plate is very hard and thick it may be softened by immersing it in warm water for some minutes. If the nail is very short, we should wait some days for it to grow.

Do you know how to send the collected material obtained by clipping to the Clinical Pathology Department?

Place the fragment in an empty vial, envelope or plastic bag. Some authors suggest placing it in formol solution at 10%, but in our experience, confirmed by the opinion of some other authors, it is not necessary ².

he pathologist and the clipping

Nail clippings, as well as nail biopsy materials, are one of the most difficult materials to make histological preparations within dermatopathology. There are many methods and required attempts to make nail tissue easy to cut. Many substances that are known to soften keratin are used, but none has proven to be satisfactory, which means that many have been tested. The most widely used are KOH at 20%, aqueous solution of Tween 40 at 10% and methachrylate ³. Some pathologists advocate histological sections of nail fragment directly included in paraffin, without previous treatment and no softener. This method has been adopted by us and resulted in good histological preparations. As a routine, slides are stained with hematoxyllin-eosin and resistant PAS-diastase or PAS-CD (With Digestion), leaving two blank slides for later staining, if necessary. By microscopy, depending on the etiology, the pathologist may detect abnormalities such as parakeratosis, serous lakes, cytoid bodies, fungi (hyphae, pseudo-hyphae, arthroconidia and blastoconidia), pigments, blood, bacteria, mites, and even foreign bodies. If hyphae are affected by treatment, it may be difficult to stain with PAS-CD; therefore, we use silver methenamine technique (Grocott), which may clarify any doubts. If necessary, we can stain with melanin pigment, which in addition to defining if it is really melanin, it positions the finding on the slide, guiding the clinician about its origin – if it is on the superficial aspect of the plate, the origin is the proximal matrix and if on the ventral aspect, the origin is the distal matrix, which facilitates the choice of the most appropriate site for the biopsy.

Do you know how to interpret the histopathological report of a clipping?

Let us compare the described findings of a pathologist with the history and physical examination. The presence of parakeratosis, serous lakes, hyperkeratosis and inflammatory cells (neutrophils) may be observed in onychomycosis and psoriasis (Figures 4 and 5). Similar findings may also be found in eczemas and trauma, but without inflammatory cells.

The presence of septated and uniform hyphae invading the plate suggests infection by dermatophytic agents (Figure 6). Hyphae with thick and tortuous walls may represent non-dermatophytic fungi and, in these cases, it is recommendable to make new culture, instructing the lab to use more appropriate medium to cultivate non-dermatophytic fungi. It may also help to explain the cases that do not respond to treatment, given that these fungi are normally resistant to antifungal agents ⁵.

Conidia on the ventral aspect of the lamina, especially if accompanied by sprouting and pseudo-hyphae, may indicate infection by Candida and, in this case, the culture is much more important to identify the genus and the species. Hyphae with degenerated aspect and isolated arthroconidia may occur as a consequence of previous exposure to antifungal agents 5. Bacterial colonies, if of large size, deserve investigation using culture and bacterioscopy. The presence of red blood cells is compatible with traumatic lesions and may be an important finding to rule out other dyschromias. The finding of melanin pigment on the nail plate confirms the diagnosis of melanonychia (Figure 7).

ARE YOU FAMILIAR WITH THESE DATA?

Signs present in onychomycosis, such as onycholysis, leuchonychia, melanonychia and total dystrophy may also be seen in other nail diseases, such as psoriasis, lichen planus, pharmacodermia and traumatic affections. It is estimated that onychomycosis represents about 50% of onychopathies. The diagnosis of these infections in clinical practice should be simple but it is not for different reasons, such as: inappropriate collection, presence of contaminants, inexperience of technicians in the preparation and identification of fungi, and liberal use of OTC medication by patients. The repetition of the exams is not always possible for social-economic reasons. Treatment of onychomycosis may be long and expensive and it may be associated with adverse events and interaction with other drugs, reason why it should be indicated only when there is confirmed diagnosis ⁶. There are few studies of onychopathy diagnosis made by use of clipping and not much familiarity of the pathologists with the topic. In turn, dermatologists are not used to make this exam in the office and have difficulty to interpret the histopathological report. The histological technique may also be impaired by inappropriate collection and insufficient size of the sample ⁷. However, as we have seen, it is an easy to perform, low cost and quick response test when compared to the time it takes to receive fungi culture results. It is a non-painful test to the patient, has no sequels and high sensitivity. It may be helpful to select the site to be biopsied in case of melanonychia and it is very important in the assessment of diseases such as psoriasis and lichen planus, when these manifestations are exclusively on the nails. Histopathological analysis of clipping is not a replacement for culture, which remains as the gold standard for diagnosis of onychomycosis owing to its high specificity. ⁸

The present paper intended to talk about a not very well known diagnostic method to dermatologists and pathologists, so that they feel encouraged to perform it and can be better fit to face the clinical challenge of making the diagnosis of nail diseases.

REFERENCES

Mailing Address:
José Fillus Neto
Rua Voluntários da Pátria, 475, 6º andar, sala
605 - Ed. Asa Centro
80020 926 Curitiba PR
Tel./fax: 55 (41) 323524
E-mail:jfn@netpar.com.br

How to cite this article: Fillus Neto J, Tchornobay AM. Como o clipping pode auxiliar o dermatologista. An Bras Dematol. 2009;84(2):173-6.