No evidence of Borrelia in cutaneous infiltrates of B-cell lymphomas with a highly sensitive, semi-nested real-time polymerase chain reaction targeting the 5S-23S intergenic spacer region.

BACKGROUND: The role of Borrelia in the development of skin lymphomas has been under discussion for decades. A similar association has been shown for Helicobacter pylori and gastric lymphomas (MALT type). Nevertheless, few molecular studies investigated Borrelia in skin lymphomas and the results are controversial. METHODS: We analysed 46 formalin-fixed, paraffin-embedded skin specimens of clincopathologically confirmed B-cell lymphomas (15 marginal zone lymphomas; 20 follicular lymphomas; three diffuse large B-cell lymphomas; eight secondary cutaneous infiltrates) taken from 36 patients from Northern Germany, an endemic area for Borrelia. Fifteen pseudolymphomatous lesions of cutaneous Borreliosis served as the control. Both groups were examined with a real-time (rt) PCR and a semi-nested PCR targeting the 5S-23S intergenic spacer region (IGS). A multiplex PCR was used to investigate B-cell clonality in all lymphomatous infiltrates (Biomed Primers). RESULTS: With both assays no Borrelia burgdorferi-specific DNA was identified in any of the B-cell lymphomas, while all 15 Borreliosis specimens gave a positive PCR result in the semi-nested PCR protocol, 12 were also positive in the rt PCR (P < 0.01). All B-cell lymphomas showed monoclonal IgH-Rearrangement. Analysis of cutaneous B-cell lymphomas from available studies including ours (n = 334) reveals an odds ratio <1. CONCLUSION: While some previous studies suggested an association between B. burgdorferi and the development of cutaneous B-cell lymphomas in endemic areas, we were unable to confirm this in our patients, despite a highly sensitive Borrelia PCR assay. Our results including meta-analysis of previous studies question the need for antibiotic therapy in patients with cutaneous B-cell lymphomas.

[Skin biopsy of inflammatory skin diseases in childhood-when is it reasonable?] / Hautbiopsie bei entzündlichen Hauterkrankungen im Kindesalter ­ wann ist sie sinnvoll?

Compared with adulthood, inflammatory skin diseases are relatively rarely biopsied in children. Apart from the invasiveness of the procedure, the required local anesthesia, and the risks of infection and scarring, the psychological trauma of the operation has a higher impact in childhood. If a biopsy is performed, expectations towards the dermatopathology report are high. However, the evaluation of biopsies taken from children is challenging for the dermatopathologist: on the one hand, because the biopsies are often tiny or just superficial shaves and, on the other hand, because criteria for evaluation have mostly been developed from findings in adult biopsy specimens. In children, the immune system is still in the process of maturation and, therefore, infiltrates in the skin may look different from those seen in adults; however, knowledge about that is very limited to date. Moreover, numerous rare genodermatoses may manifest themselves first in childhood and need to be considered in the differential diagnosis while experience with them is often limited. Starting from the clinical presentation, this article presents histopathological features of possible differential diagnoses in order to demonstrate the value or necessity of a skin biopsy in a pediatric patient. In addition, communication with parents and child, methods of local anesthesia and biopsy techniques will be considered.

[Relevance of biopsies for the diagnostics of infectious skin diseases]. / Stellenwert der Biopsie in der Diagnostik von Infektionserkrankungen der Haut.

Pathogen-related skin infections are a common problem in the dermatological practice. Apart from culturing and serological detection methods, a skin biopsy is a possible diagnostic procedure, especially when the clinical picture is unspecific and other non-infectious skin diseases are considered as possible differential diagnoses. Some organisms can already be detected by routine staining methods (hematoxylin & eosin, e. g., yeasts, Leishmania), for others numerous histochemical and immunohistochemical stains are available, e. g. periodic acid-Schiff reaction (PAS) and Grocott for hyphae and spores, Ziehl-Neelson and Fite-Faraco for Mycobacteria or specific antibodies for Treponema pallidum or herpesviruses. In other instances, an infectious disease may not be diagnosed with certainty in a histological section but the pattern of inflammatory infiltrates is highly suggestive of an infectious cause. Based on such reaction patterns, the dermatopathologist can advise the clinician to perform cultures or serological investigations or additional molecular biological techniques can be applied to the biopsy specimen in order to identify the pathogens. This article presents skin infections with their histopathological features and highlights diseases that can be diagnosed with certainty in a biopsy and those in which a biopsy is helpful to exclude differential diagnoses or to perform molecular diagnostics on the specimen.

[Differential diagnosis of panniculitides]. / Differenzialdiagnostik der Pannikulitis.

Panniculitides are diseases of the subcutaneous tissue with heterogeneous etiology. They may develop consequent to infections, as a reaction to drugs, after thermal injury, as part of autoimmune diseases, in metabolic disorders or due to infectious organisms. The clinical presentation with subcutaneous nodules is often nonspecific. Moreover, the differentiation from vasculitides of medium-sized vessels can be clinically challenging. Microscopic examination of biopsy specimens is of high importance in the differential diagnosis of panniculitides. Histopathologically, panniculitides can be classified according to the predominantly infiltrated area in septal and lobular panniculitides and they can be separated from vasculitides of medium-sized vessels. Diagnostic difficulties arise from inadequate biopsy specimens and from lack of clinicopathological correlation. This article summarizes diagnostic criteria of frequent and clinically important panniculitides.

Species identification of dermatophytes in paraffin-embedded biopsies with a new polymerase chain reaction assay targeting the internal transcribed spacer 2 region and comparison with histopathological features.

BACKGROUND: Dermatophytosis is a very common skin infection with a broad clinical spectrum. Biopsies are often used to confirm the diagnosis, especially when the clinical presentation is unusual. Not uncommonly, organisms are hard to find even with periodic acid-Schiff stains. Polymerase chain reaction (PCR) for dermatophytes can be used in such cases. OBJECTIVES: To test a new PCR assay allowing species identification of dermatophytes on paraffin-embedded biopsies, and to reassess histopathological criteria for diagnosis of dermatophytosis. METHODS: In total, 121 biopsies of 92 patients with clinical suspicion of tinea were included. In 42 samples the clinical diagnosis had been confirmed histopathologically, and in 79 no fungal elements had been identified. PCRs targeting the internal transcribed spacer (ITS)2 region of dermatophytes were performed on the biopsies with subsequent sequencing. Sections were reassessed for the presence/absence of hyphae/spores, pattern and composition of infiltrate, and epidermal/follicular changes. Patient charts were reviewed for clinical data. RESULTS: The new ITS2 PCR assay detected 94% of the dermatophyte infections (compared with 79% identified by microscopy). Trichophyton rubrum was the dominant species (89%), and other species identified were Trichophyton verrucosum (2%), Microsporum canis (4%), Epidermophyton floccosum (2%) and Trichophyton interdigitale (4%). In particular, infections with T. interdigitale and manifestations with prominent spongiosis were not diagnosed histologically. Intracorneal neutrophils, which have been emphasized as a histopathological clue to dermatophytosis, were present in only 46% of PCR-positive samples. CONCLUSIONS: Molecular species identification of dermatophytes via ITS2 PCR can easily be implemented in a routine dermatopathology setting. It is fast and highly specific and improves the sensitivity of histopathological diagnosis of dermatophytosis.

Fast, sensitive and specific diagnosis of infections with Leishmania spp. in formalin-fixed, paraffin-embedded skin biopsies by cytochrome b polymerase chain reaction.

BACKGROUND: Northern spread of sandflies and Leishmania spp. has been observed in Europe. Diagnosis can be difficult owing to the various clinical manifestations. Species identification is important for patient management and therapy. Molecular diagnostics is increasingly used for pan-Leishmania detection but species identification remains challenging in formalin-fixed material. OBJECTIVES: To apply cytochrome b (cytb) polymerase chain reaction (PCR) and sequencing for identification of Leishmania species on formalin-fixed, paraffin-embedded (FFPE) skin biopsies; and to identify species-specific histological patterns. METHODS: Sixty-nine biopsies (48 patients) diagnosed with leishmaniasis based on the presence of amastigotes in the tissue (n = 41) or granulomatous infiltrates with positive pan-Leishmania real-time PCR (n = 28) were analysed with cytb PCR, sequencing and phylogenetic analysis. Histological sections were analysed; epidemiological data were collected. RESULTS: Cytb PCR identified Leishmania in all specimens: L. infantum (79%), L. major (8%), L. panamensis (4%), L. tropica (4%), L. killicki (2%) and L. aethiopica (2%). Of the detected species 95% were endemic to the country in which the infection was acquired. Amastigotes were found in 59%. Infiltrates were mainly tuberculoid granulomatous (65%), interstitial (15%) and sarcoidal (10%). Pseudolymphomatous features and pseudocarcinomatous hyperplasia were more common in L. major infections than in L. infantum (P < 0·01). CONCLUSIONS: Cytb PCR and sequencing is a fast, reliable and sensitive assay for identification of Leishmania spp. in FFPE biopsies. Leishmania infantum is the main cause of cutaneous leishmaniasis in Germany. Tuberculoid granulomas, other types of granulomas and pseudolymphomatous infiltrates may be encountered; the latter being indicative of infection with L. major.

[Cutaneous granulomatous reactions at botulinum neurotoxin A injection sites: First manifestation of systemic sarcoidosis]. / Granulomatöse Hautveränderungen an Botulinumtoxin-A-Injektionsstellen : Erstmanifestation einer systemischen Sarkoidose.

CASE REPORT: We report a case of granulomatous skin reaction after injection of an unknown botulinum neurotoxin (BoNT) A preparation. Four years after occurrence of skin lesions, systemic sarcoidosis became manifest. CONCLUSION: In addition to injection trauma, the BoNT A preparation may have acted as an immunogenic stimulus leading to cutaneous manifestation of sarcoidosis.

[Differential diagnostics of sebaceous tumors]. / Differenzialdiagnostik der Talgdrüsentumoren.

Sebaceous tumors are epithelial tumors with a differentiation towards sebaceous adnexal structures of the skin. They imitate the epithelial cells of mature sebaceous glands, sebaceous ducts, immature (embryonic) sebaceous structures or sebaceous glands that are not stimulated by hormones (mantle structures). This article explains the classification of sebaceous tumors on the basis of the normal histology of sebaceous glands. Clinical and histopathological criteria are given for the most important sebaceous tumors. The differential diagnosis of sebaceoma, sebaceous adenoma and various types of sebaceous carcinoma is emphasized. The importance of a specific diagnosis of adnexal tumors is demonstrated by tumor-associated syndromes with involvement of other organs (e.g., Muir-Torre syndrome and Birt-Hogg-Dubé syndrome). Furthermore, conceptional controversies, problems in differential diagnosis and the impact of immunohistochemical staining in the assessment of sebaceous tumors are considered.

[Fundamentals of cutaneous adnexal tumors]. / Basiswissen Hautadnextumoren.

Adnexal tumors of the skin are epithelial skin tumors with differentiation towards the adnexal epithelial structures of the skin, namely, hair follicle, sebaceous gland, apocrine gland, and eccrine gland. Adnexal tumors include hamartomas, benign and malignant neoplasms, and hyperplasias. The specific diagnosis of adnexal tumors is important because some lesions such as sebaceous neoplasms, cylindromas, or fibrofolliculomas are herald lesions of hereditary tumor syndromes (e.g., Muir-Torre syndrome, familial cylindromatosis, Birt-Hogg-Dubé syndrome). In this article, the classification of adnexal tumors of the skin is explained on the basis of embryology and histology and the main features of tumor-associated syndromes are summarized. Moreover, some conceptual controversies and problems in differential diagnosis of cutaneous adnexal tumors are discussed.

Genotyping of Borrelia from formalin-fixed paraffin-embedded skin biopsies of cutaneous borreliosis and tick bite reactions by assays targeting the intergenic spacer region, ospA and ospC genes.

BACKGROUND: Lyme borreliosis has a broad spectrum of clinical presentations involving the skin, joints and nervous system. The variable manifestations have been attributed to different Borrelia genospecies but genotyping required culture or fresh tissue. However, in dermatology practice, formalin-fixed paraffin-embedded biopsies are used for dermatopathological examination. Polymerase chain reaction (PCR) for Borrelia burgdorferi sensu latu has been established on such specimens, but studies attempting genotyping of subspecies or strains are lacking. OBJECTIVES: To adapt PCR assays for genotyping of Borrelia using paraffin-embedded biopsies, to identify Borrelia genospecies and to compare clinicopathological features of different genospecies. METHODS: Eighty-two paraffin-embedded biopsies from 68 patients, with erythema migrans, acrodermatitis chronica atrophicans, lymphocytoma cutis or tick bite reactions, were studied with assays targeting the intergenic spacer (IGS), ospA and ospC, followed by sequencing and phylogenetic analysis. Clinicopathological data were analysed comparing different Borrelia genospecies. RESULTS: Genotyping by IGS, ospA and ospC was successful in 85% of patients (91% B. afzelii, 7% B. garinii, 2% B. bavariensis). ospA serotyping identified type 2 (90%), type 3 (8%) and type 4 (2%). ospC-PCR was positive in 40% of the patients revealing 12 different groups, noninvasive forms being seen only in tick bite reactions and erythema migrans. No major clinicopathological differences could be identified between the genospecies, but neural inflammation and arthralgia were seen more often in lesions caused by invasive ospC strains. CONCLUSIONS: Genotyping of Borrelia can be easily implemented in a routine dermatopathology setting, especially as a fast method to confirm early cutaneous borreliosis. Genotyping could also enable earlier treatment of patients infected with invasive strains.

[Fundamentals of histology]. / Basiswissen Histologie.

Very commonly skin biopsies are crucial for the correct diagnosis of patients with skin diseases. The dermatopathology report is an important tool for communication between the clinical dermatologist and the dermatopathologist. This article explains the method of histopathological assessment of skin biopsies and summarizes the most important criteria for diagnosis of common skin diseases. The application of additional histochemical, immunohistochemical, and molecular biological methods is elucidated. The clinical dermatologist requires basic knowledge of dermatopathology to decide on the appropriate biopsy procedure. It also enables him to understand the dermatopathology report and to identify settings in which clinicopathological correlation is required.

Benign mucinous metaplasia of the genital mucosa: histomorphological and immunohistochemical features and criteria for differentiation from extramammary Paget disease.

BACKGROUND: Benign mucinous metaplasia of the genitalia (BMM) is a rare condition typified by cells with foamy mucinous cytoplasm. Differential diagnoses include extramammary Paget disease (PD) and human papillomavirus (HPV)-induced vulval intraepithelial neoplasia (VIN) with mucinous differentiation. OBJECTIVES: To characterize histopathological and immunohistochemical features of BMM and to forge criteria for differentiation from PD and VIN with mucinous differentiation. METHODS: Eight biopsy specimens of BMM were stained with haematoxylin and eosin, periodic acid-Schiff and alcian blue, and for cytokeratin (CK) 7, CK10, CK14, CK20, carcinoembryonic antigen (CEA), epithelial membrane antigen (EMA), S100, gross cystic disease fluid protein-15 (GCDFP-15), lysozyme and Ki67 and compared with PD. Polymerase chain reaction was performed in order to identify HPV-specific DNA. RESULTS: BMM showed mucin deposition in superficial epithelial layers ranging from numerous large goblet cells to subtle deposits. The epithelium often showed polygonal (squamoid) or cuboidal differentiation while columnar differentiation was an inconsistent feature. A band-like inflammatory infiltrate was consistently present. Metaplastic epithelium consistently expressed CK7, CEA and EMA either in the entire epithelium or in a superficial band, while CK14, CK10, GCDFP-15 and lysozyme were largely not expressed, and staining for CK20 and S100 was negative. Comparison with PD demonstrated similar staining characteristics, but in a scattered pattern of mucinous cells within preserved squamous epithelium and not in a band-like pattern as in BMM. Nuclear pleomorphism and Ki67-positive mucinous cells in superficial epithelial layers were seen only in PD; GCDFP-15 and/or lysozyme were expressed in the majority of cases of PD. No evidence of HPV-specific DNA was found in BMM. CONCLUSIONS: The spectrum of changes in BMM is distinctive, and BMM can be differentiated with surety from both PD and VIN with mucinous differentiation.