A possible association of Borrelia burgdorferi with localized scleroderma is currently the focus of intense research and discussion. Skin biopsies from 30 patients with localized scleroderma (28 of the plaque type/morphea; two linear scleroderma) were analyzed for the presence of Borrelia burgdorferi using three different polymerase chain reaction systems for amplification of segments of borrelial genes. Formalin-fixed, paraffin-embedded biopsies of 14 patients and fresh-frozen, cryo-conserved biopsies of 16 patients with localized scleroderma were obtained. Lesions of all patients showed clear signs of scleroderma and disease progression at the time of biopsy. Fresh-frozen as well as formalin-fixed biopsies from patients with erythema migrans or acrodermatitis chronica atrophicans were used as positive controls. With all three polymerase chain reaction systems, borrelial DNA was detected in none of the 30 specimens of localized scleroderma. In contrast, with one polymerase chain reaction system, Borrelia burgdorferi-specific DNA was found in 24 of 27 frozen biopsies from patients with erythema migrans and in all 5 analyzed frozen biopsies of patients with acrodermatitis chronica atrophicans. In approximately half of the paraffin-embedded biopsies from patients with erythema migrans (nine of 23) and acrodermatitis chronica atrophicans (13 of 27), Borrelia burgdorferi-specific DNA was identified. These results question the association of localized scleroderma with known subtypes of Borrelia burgdorferi.

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Background: Herpes simplex virus infections are well known complications of various dermatoses and have also been reported in acantholytic diseases like pemphigus vulgaris or Darier’s and Hailey-Hailey diseases. In pemphigus vulgaris, herpes simplex virus infection is considered to be rare and difficult to rule out clinically. Objective: We report on 3 patients suffering from pemphigus vulgaris with exacerbation especially of lesions of the oral mucosa. Methods and Results: While conventional techniques failed to unequivocally support a suspected herpetic infection, herpes simplex virus-specific DNA was detected by polymerase chain reaction (PCR) in cytological swabs taken from oral erosions of all 3 patients. Conclusion: Herpetic infection should be considered in pemphigus vulgaris with lack of improvement under adequate immunosuppressive therapy. In addition, herpes simplex virus infection might to able to induce acute exacerbation of oral pemphigus. PCR can be useful for a highly sensitive and rapid molecular detection of herpes simplex virus.

The identification of circulating tumor cells in the peripheral blood of patients with malignant melanoma by detection of melanoma associated protein transcripts using the reverse transcriptase polymerase chain reaction (RT-PCR) technique has been introduced as a noninvasive and sensitive technique for early detection of tumor progression and metastatic disease. An alternative approach is the analysis of S-100 protein in the serum of melanoma patients by a luminoimmunometric assay (LIA). In this study, the sensitivities of RT-PCR and LIA were compared. Seventy-seven blood samples of 59 melanoma patients were analyzed for tyrosinase, Melan-A/MART-1, MAGE-3, gp100, and p97 expression by multimarker RT-PCR; 540 serum samples of 352 melanoma patients were analyzed for S-100 protein concentration by LIA. In stage III 23.8% and in stage IV 37.5% of the samples were positive for at least one marker in multimarker RT-PCR, versus 8.1% and 48.1% of elevated S-100 levels analyzed by LIA, respectively. In a direct comparison, 31 identical samples were analyzed by multimarker RT-PCR and by S-100 LIA. In stage III 18.2% and in stage IV 45% of the samples were positive by multimarker RT-PCR versus 45.5% and 80% by S-100 LIA, respectively. S-100 LIA was more sensitive in detection of metastatic disease in melanoma patients than multimarker RT-PCR and should be evaluated in further studies. RT-PCR might be more useful in the analysis of micrometastases in anatomic compartments other than peripheral blood.

SUMMARYMelanoma-speci®c cytotoxic T lymphocytes (CTL) can be generated from peripheral blood lymphocytes (PBL) by mixed lymphocyte±tumour cell cultures. Analysis of CTL precursor frequencies in peripheral blood of melanoma patients is generally used for immunomonitoring purposes to evaluate vaccination ef®cacy. At present, it is unclear whether PBL-derived CTL generated in vitro are indicative of an anti-tumour immune response in vivo. Three tumour-speci®c human leucocyte antigen (HLA)-B/C-restricted CTL clones were derived from peripheral blood of a melanoma patient immunized with interleukin-7 (IL-7) gene-modi®ed tumour cells. CTL clones differing in their T-cell receptor-c (TCRc) rearrangement produced interferon-c, IL-4 and/or IL-10. On the basis of their unique TCRc gene rearrangements clone-speci®c primers were generated for detection of clone-speci®c DNA by polymerase chain reaction. One CTL clone (E5) of the three was found to be selectively expanded in one of seven metastases obtained at autopsy, as determined by Southern blot hybridization. However, the presence of E5 in only one of seven metastases at death indicates that the in vivo accumulation of the speci®c CTL clone was not suf®cient to contain tumour progression. Nevertheless, our data support the proposition that analysis of anti-tumour activity of PBL-derived CTLs may re¯ect an anti-tumour immune response in vivo.

A 51-year-old human immunodeficiency virus (HIV)-positive male patient (CDC stage 3C) had had a painful nodule on his external ankle joint for 10 months. A biopsy suggested bacillary angiomatosis, but Kaposi's sarcoma could not be excluded. Rods were detectable in lesional skin by a Warthin-Starry stain. A 298 base pair (bp) gene fragment specific for Bartonella species was amplified from lesional skin and direct nucleotide sequence analysis of the amplification product clearly identified Bartonella quintana. Kaposi's sarcoma-associated herpes virus specific DNA was not amplifiable by polymerase chain reaction (PCR) in our patient, suggesting that the lesion represented bacillary angiomatosis alone, despite clinical and histopathological features which suggested the coexistence of bacillary angiomatosis and Kaposi's sarcoma. The lesion regressed after erythromycin was prescribed. However, 4 and 9 weeks after initiation of therapy, PCR still yielded a positive result in material obtained by a swab. After complete healing, following 12 weeks of antibiotic therapy, PCR became consistently negative. The optimal length of antibiotic treatment in HIV-positive patients with bacillary angiomatosis is not yet known and inadequate therapy may be followed by disseminated disease and a fatal outcome. PCR-based monitoring of the success of treatment is valuable for determining the duration of treatment resulting in a cure.

A 49-year-old man suffered from Hailey-Hailey disease for several years. The patient presented with an acute exacerbation of the disease, which did not respond to oral treatment with high doses of glucocorticosteroids. A skin biopsy was taken and the histological examination indicated a viral infection. Herpes simplex virus was confirmed by electron microscopy (negative staining) and polymerase chain reaction (PCR). The PCR presents a sensitive and effective molecular-biological method for the diagnosis of viral infections.