BCC and Immunocryosurgery scar differentiation through computational resolution-enhanced OCT images and skin optical attenuation: A proof-of-concept study.

Monitoring medical therapy remains a challenging task across all non-surgical skin cancer treatment modalities. In addition, confirmation of residual tumours after treatment is essential for the early detection of potential relapses. Optical coherence tomography (OCT), a non-invasive method for real-time cross-sectional imaging of living tissue, is a promising imaging approach for assessing relatively flat, near-surface skin lesions, such as those that occur in most basal cell carcinomas (BCCs), at the time of diagnosis. However, the skin's inherent property of strong light scattering impedes the implementation of OCT in these cases due to the poor image quality. Furthermore, translating OCT's optical parameters into practical use in routine clinical settings is complicated due to substantial observer subjectivity. In this retrospective pilot study, we developed a workflow based on the upscale of the OCT images resolution using a deep generative adversarial network and the estimation of the skin optical attenuation coefficient. At the site of immunocryosurgery-treated BCC, the proposed methodology can extract optical parameters and discriminate objectively between tumour foci and scar tissue.

Immunocryosurgery is a safe and feasible treatment for basal cell carcinoma and Bowen disease in renal transplant recipients.

Keratinocyte skin carcinomas (squamous cell carcinoma, basal cell carcinoma [BCC], Bowen disease [BD]) inflict significant morbidity and constitute a treatment challenge in renal transplant recipients (RTR). Immunocryosurgery has shown efficacy >95% in the treatment of BCC and BD in immunocompetent patients. The present study evaluated the safety, feasibility and efficacy, of immunocryosurgery in the treatment of BCC and BD in a series of RTR. During a 3-year period, biopsy-confirmed cases of BCC and BD were treated with a standard immunocryosurgery cycle (5 weeks daily imiquimod and a session of cryosurgery at day 14). Safety was evaluated by comparing graft function markers between immunocryosurgery treated RTR patients and matched controls. Ten BCC (8 nodular, 1 basosquamous, 1 superficial; diameter 6-14 mm; mean 9.2 mm) and nine BD disease lesions in nine patients (7 men, 2 women; age range: 54-70 years, mean: 62.1 years) were treated with immunocryosurgery and followed-up for two to 5 years. Five BCC were located on the "H area" of the face. No patient showed clinical or laboratory signs of transplant dysfunction during treatment or follow-up. Seven out of 10 BCC lesions cleared completely after one 5-week immunocryosurgery cycle, two cleared after repeat and intensified treatment cycles and one responded only partially (clearance rate: 90%). Seven out of nine BD lesions cleared after one 5-week immunocryosurgery cycle and one lesion after two cycles (clearance rate: 88.9%). In conclusion, immunocryosurgery is a safe, feasible and effective minimally invasive treatment alternative to standard surgical modalities for BCC and BD in RTR.

Spirochetal uveitis: Spectrum of clinical manifestations, diagnostic and therapeutic approach, final outcome and epidemiological data.

PURPOSE: Analysis of cases with spirochetal uveitis related to spirochetes in a tertiary referral academic center. METHODS: Retrospective study of patients diagnosed with uveitis attributed to Treponema pallidum, Leptospira spp. and Borrelia burgdorferi from June 1991 until December 2019. RESULTS: A total of 57 cases of spirochetal uveitis (22 patients with T. pallidum, 26 with Leptospira spp., and 9 with B. burgdorferi) that consisted 1% of the overall number of uveitics were recorded. All these cases presented with a wide spectrum of clinical presentations (anterior uveitis, posterior uveitis, panuveitis, vasculitis, papillitis, and in some rare cases concomitant posterior scleritis). The treatment included mainly penicillin or doxycycline, while corticosteroids were administered systematically in some cases with Borrelia or Leptospira infection. The final visual outcome was favorable (> 6/10 in Snellen visual acuity) in approximately 76% of our patients. CONCLUSION: Despite being rare, spirochetal uveitis can be detrimental for the vision and must always be included in the differential diagnosis.

The lipid profile of three Malassezia species assessed by Raman spectroscopy and discriminant analysis.

Malassezia yeasts constitute the major eukaryotic cutaneous flora of homoeothermic vertebrates. These lipophilic yeasts are able to cause, trigger, or aggravate common skin diseases under favorable conditions. Species identification and subspecies differentiation is currently based on morphological characteristics, lipid assimilation profile, and molecular tests. Mass spectrometry has been also reported as a reliable, yet costly and labor-intensive, method to classify Malassezia yeasts. Here, we introduce Raman spectroscopy as a new molecular technique able to differentiate three phylogenetically close Malassezia species (M.globosa, M.pachydermatis, and M.sympodialis) by examining their lipid metabolic profile. Using Raman spectroscopy, lipid fingerprints of Malassezia cultures on Leeming-Notman agar, were analyzed by spectral bands assignment and partial least squares discriminant analysis. Our results demonstrate differential utilization of lipid supplements among these three species and the ability of Raman spectroscopy to rapidly and accurately discriminate them by predictive modelling.

Late fusion of deep and shallow features to improve discrimination of actinic keratosis from normal skin using clinical photography.

BACKGROUND: Actinic keratosis (AK) is a common premalignant skin lesion that can potentially progress to squamous cell carcinoma. Appropriate long-term management of AK requires close patient monitoring in addition to therapeutic interventions. Computer-aided diagnostic systems based on clinical photography might evolve in the future into valuable adjuncts to AK patient management. The present study proposes a late fusion approach of color-texture features (shallow features) and deep features extracted from pre-trained convolutional neural networks (CNN) to boost AK detection accuracy on clinical photographs. MATERIALS AND METHODS: System uses a sliding rectangular window of 50 × 50 pixels and a classifier that assigns the window region to either the AK or the healthy skin class. 6010 and 13 915 cropped regions of interest (ROI) of 50 × 50 pixels of AK and healthy skin, respectively, from 22 patients were used for system implementation. Different support vector machine (SVM) classifiers employing shallow or deep features and their late fusion using the max rule at decision level were compared with the McNemar test and Yule's Q-statistic. RESULTS: Support vector machine classifiers based on deep and shallow features exhibited overall competitive performances with complementary improvements in detection accuracy. Late fusion yielded significant improvement (6%) in both sensitivity (87%) and specificity (86%) compared to single classifier performance. CONCLUSION: The parallel improvement of sensitivity and specificity is encouraging, demonstrating the potential use of our system in evaluating AK burden. The latter might be of value in future clinical studies for the comparison of field-directed treatment interventions.

Antifungal activity of selected Malassezia indolic compounds detected in culture.

BACKGROUND: Malassezia yeasts produce bioactive indolic substances when grown on L-tryptophan agar. A panel of these substances was tested against commensal and opportunistic fungi, the Minimum Inhibitory Concentration (MIC) was determined and the potential for in loco antifungal activity on the skin was assessed. MATERIALS AND METHODS: Eight indoles were included (malassezin, pityriacitrin, indirubin, indolo[3,2-b]carbazole, 6-formylindolo[3,2-b]carbazole, tryptanthrin, 6-hydroxymethylindolo[3,2-b]carbazole and 6-methylindolo[3,2-b]carbazole) and were tested against 40 fungal strains [yeasts: Malassezia spp.(N = 9); Cryptococcus spp.(N = 10); Candida spp.(N = 7); Yarrowia lipolytica(N = 1); Exophialla dermatitidis (N = 2); moulds: Aspergillus spp.(N = 7); Fusarium spp.(N = 2); Rhizopus oryzae(N = 2)]. The concentration of 5/8 of the tested indoles on diseased skin was calculated from published data. Kruskal-Wallis and Mann-Whitney U tests were employed for group susceptibility evaluation in 33 strains. RESULTS: The MIC range was 0.125-32 µg/mL, and the median log2 MIC was four. Indirubin was the most potent antifungal agent and differed significantly from the others. The highest median MIC was found for FICZ. Malassezia with Candida strains were more susceptible compared to Cryptococcus and Aspergillus, and this inhibitory activity was predicted to be valid also on human skin. CONCLUSIONS: Malassezia yeasts produce indolic species that inhibit an array of clinically significant yeasts and moulds.

Variation of cultured skin microbiota in mothers and their infants during the first year postpartum.

BACKGROUND/OBJECTIVES: The establishment of newborn skin flora depends on the ongoing skin maturation and the existence of potential microbial colonizers within the environment of the infant during a period of intense mother-infant physical interaction. This longitudinal study assessed culturable skin bacteria in the mother-infant dyad during the first year of life. METHODS: A total of 17 mother-infant dyads were swabbed within 24 hours postpartum and at 3, 6, 9, and 12 months. Skin swabbing was performed on two anatomical areas per individual (mothers: chest-abdomen; infants: forehead-buttocks) and were incubated in five different solid culture media to optimize yield. Isolated bacterial species were identified to genus or species level using the API system (BioMeriéux, Marcy l'Etoile, France). RESULTS: A total of 444 microbial strains were isolated belonging to 22 genera: 6 "frequent" (isolated from > 5% samples: S aureus, Proteus, Klebsiella, Pseudomonas, Enterobacter, and Enterococcus) and 16 "infrequent." Isolated genera per individual peaked at 6 months postpartum for mothers and infants (P < 0.05). Enterobacter, Enterococcus, Klebsiella, and Pseudomonas isolation rates varied significantly as a function of sampling time contrary to the rather constant isolation rates of Proteus and S aureus. The rates of concordant isolation of the same microbial species within the mother-infant dyad tended to drop from birth to the end of the first year postpartum. CONCLUSIONS: Distinct variations in the isolation rates of skin commensals from specific anatomical sites of the mother-infant dyad indicate bidirectional microbial transmission. Increasing skin flora individuality of the growing infant was recorded, manifested by declining rates of concordant isolation of the same microbial species from mother and her infant.

Nail Raman spectroscopy: A promising method for the diagnosis of onychomycosis. An ex vivo pilot study.

Trichophyton rubrum and Candida species comprise the majority of onychomycosis pathogens. The aim of this study was to evaluate Raman spectroscopy for the differentiation between healthy and either T. rubrum or Candida infected nails. Raman measurements were performed on clippings (N = 52) infected either by T. rubrum (N = 12) or Candida species (N = 14; C. parapsilosis (sensu lato): N = 11, C. glabrata: N = 1, C. albicans: N = 2) with healthy nails (N = 26) used as controls. Systematic spectral differences were observed in the 500-520 cm-1 band region, attributable to a diverting imprint of the disulfide stretching of cystine and cysteine residues among samples. Particularly, Candida infected nails demonstrated a shoulder at 519 cm-1, corresponding to the signal of the less stable gauche-gauche-trans conformation of the disulfide bond. Two additional bands at 619 and 648 cm-1, corresponding to the C-S stretching vibration, were more evident in the T. rubrum infected nails. Finally, a Raman band at 1550 cm-1, attributable to amide II and tryptophan (Trp) content, was undetectable in Candida infected nails. Using principal component analysis (PCA), efficient differentiation of healthy, T. rubrum and Candida species infected nails was achieved. Soft independent modeling of class analogy (SIMCA) and partial least squares-discriminant analysis (PLS-DA) were further applied to generate diagnostic algorithms for the classification of Raman spectra. Both techniques succeeded in modeling clinical nail samples in three groups according to their mycological categories. Raman spectroscopy is a promising method for the differentiation of healthy vs. diseased nails, including efficient differentiation between onychomycosis caused by T. rubrum and Candida species.

Malassezia ecology, pathophysiology, and treatment.

Malassezia are lipid dependent basidiomycetous yeasts that inhabit the skin and mucosa of humans and other warm-blooded animals, and are a major component of the skin microbiome. They occur as skin commensals, but are also associated with various skin disorders and bloodstream infections. The genus currently comprises 17 species and has recently been assigned its own class, Malasseziomycetes. Importantly, multiple Malassezia species and/or genotypes may cause unique or similar pathologies and vary in their antifungal susceptibility. In addition to culture-based approaches, culture-independent methods have added to our understanding of Malassezia presence and abundance and their relationship to pathogenicity. Moreover, these novel approaches have suggested a much wider-spread presence, including other human body parts and even other ecosystems, but their role in these arenas requires further clarification. With recent successful transformation and genetic engineering of Malassezia, the role of specific genes in pathogenesis can now be studied. We suggest that characterizing the metabolic impact of Malassezia communities rather than species identification is key in elucidation of pathophysiological associations. Finally, the increasing availability of genome sequences may provide key information aiding faster diagnostics, and understanding of the biochemical mechanisms for Malassezia skin adaptation and the design of future drugs.

Higher Serum Ferritin Levels Correlate with an Increased Risk of Cutaneous Morbidity in Adult Patients with ß-Thalassemia: A Single-Center Retrospective Study.

Disturbed iron homeostasis characterizes ß-thalassemia and increases its morbidity. Our aim was to retrospectively associate ß-thalassemia disease characteristics with treatment-requiring skin conditions. The files of adult ß-thalassemia (including sickle ß-thalassemia) patients were screened over a 10-year period for treatment-requiring skin disease episodes and their correlation with hematologic diagnoses and epidemiological and serological characteristics. Seventy-eight patients were identified, and 7 (9%) developed at least one relevant episode including cutaneous small-vessel vasculitis (CSVV), urticaria, and leg ulcers. Average ferritin serum level correlated significantly with development of a dermatosis (2,034 ± 799 µg/l in cases vs. 920 ± 907 µg/l in the overall population; p = 0.001, ANOVA). This difference relied exclusively on the high ferritin levels observed in patients with 'generalized' dermatoses (urticaria and CSVV: 3,860 ± 1,220 µg/l) as opposed to values within the normal range in the case of 'localized' ones (leg ulcers: 662 ± 167 µg/l). The employed iron chelation treatment influenced ferritin levels (p = 0.002, Kruskal-Wallis test) since chelation with a single agent seems to increase the risk of a skin disease (p = 0.013, likelihood ratio method). Conclusively, serum ferritin can be evaluated as risk factor for generalized dermatoses, but not for leg ulcers, in patients with the ß-thalassemia genotype. This risk can be efficiently controlled with adequate chelation.

Immunocryosurgery - an effective combinational modality for Bowen's disease.

Bowen's disease (BD) is widely treated with topical imiquimod or cryosurgery. The present single-center retrospective study reports on the application of standardized immunocryosurgery (cryosurgery during ongoing topical imiquimod) for the treatment of BD. Daily imiquimod 5% cream was applied on BD lesion and a 5 mm rim around it in 5-week treatment cycles; cryosurgery (liquid N2 , open spray; 2 cycles, 15 second each) was performed at the end of the second week of each treatment cycle. Between 1/1/2009 and 31/12/2014 21 patients (mean age ± SD: 74.4 ± 8.0 years; 12 males) with 24 lesions (mean maximum diameter ± SD: 45.8 ± 50.9 mm; range: 9-200 mm) completed the protocol. The anatomic distribution of the lesions included face/scalp (Ν = 14), neck/trunk (Ν = 6), and extremities (Ν = 4). Twenty-one out of twenty-four lesions with diameter <80 mm cleared after one immunocryosurgery cycle, while the rest three tumors (with the largest diameters: 100, 180, 200 mm) required two treatment cycles for complete response (clearance rate: 100%). After a median follow-up of 24 months (range: 6-60 months) the overall effectiveness was 91.7%: 22/24 lesions remained in sustained complete remission. With the exception of a variable degree of hypopigmentation, the cosmetic outcome was satisfactory even for extensive lesions. Immunocryosurgery, is feasible and highly efficacious minimally-invasive treatment alternative for BD.

Cryosurgery during Imiquimod (Immunocryosurgery) for Periocular Basal Cell Carcinomas: An Efficacious Minimally Invasive Treatment Alternative.

BACKGROUND/AIM: Periocular basal cell carcinomas (pBCC) remain a treatment challenge. Our aim was to retrospectively evaluate the feasibility and efficacy of immunocryosurgery for the treatment of pBCC. METHODS: Immunocryosurgery is given in 5-week cycles of daily imiquimod, with cryosurgery on day 14. Patients treated between 1/1/2008 and 31/12/2014 were included in this study. RESULTS: Immunocryosurgery was offered to 19 patients. Of these, 16 (i.e., 6 males and 10 females, average age 74.9 years, median tumor diameter 15 mm, range 5-60 mm), with 1 tumor each, were treated. Six tumors (37.5%) were relapses after surgery and 2 were of metatypical histology. All BCC were high risk for recurrence after treatment; 10 tumors had 2 risk factors for relapse, 5 had 3, and 1 had 4. The follow-up period ranged between 3 and 60 months (average 25.6 months). Of the 16 tumors treated, 14 (all with a diameter <40 mm) cleared with immunocryosurgery (total efficacy 87.5%); 7 out of 16 tumors (44%; all with a diameter ≤20 mm) cleared with 1 conventional 5-week immunocryosurgery treatment cycle. Seven additional tumors (including 2 with a diameter >20 mm) required intensified treatment schemes (of up to 10 weeks) for clearance. The 2 tumors that did not clear responded partially and were also the 2 largest ones (diameter 40 and 60 mm). Of the 14 cleared tumors, 2 relapsed during follow-up; 1 cleared with immunocryosurgery. At the last examination during follow-up, 13 out of 16 (81%) patients were in sustained clinical remission. CONCLUSIONS: For most pBCC, immunocryosurgery is a feasible and efficacious alternative to surgical excision.

An Aggregate of Four Anthrax Cases during the Dry Summer of 2011 in Epirus, Greece.

BACKGROUND: Human anthrax is currently a sporadic disease in Europe, without significant regional clustering. OBJECTIVE: To report an unexpected aggregate of anthrax cases and correlate local climatic factors with yearly anthrax admissions. METHODS: Clinical description of a geographical-temporal anthrax aggregate, correlation of disease admissions with local weather data in the period 2001-2014 and literature reports of anthrax clusters from Europe in the last 20 years. RESULTS: We identified 5 cases, all cutaneous: an unexpected aggregate of 4 cases in mid-summer 2011 (including a probable human-to-human transmission) and a sporadic case in August 2005, all in relatively dry periods (p < 0.05). Remarkably, 3/6 reports of human anthrax aggregates from Europe were observed in Balkan Peninsula countries in the year 2011. CONCLUSION: In the light of the predicted climatic change, unexpected anthrax aggregates during dry periods in southern Europe underscore the risk of future anthrax re-emergence on this continent.

Pityriazepin and other potent AhR ligands isolated from Malassezia furfur yeast.

Malassezia furfur yeast strains isolated from diseased human skin preferentially biosynthesize indole alkaloids which can be detected in the human skin and are highly potent activators of the aryl hydrocarbon receptor (AhR) and AhR-dependent gene expression. Chemical analysis of an EtOAc extract of a M. furfur strain obtained from diseased human skin and grown on l-tryptophan agar revealed several known AhR active tryptophan metabolites along with a previously unidentified compound, pityriazepin. While its structure resembled that of the known alkaloid pityriacitrin, the comprised pyridine ring had been transformed into an azepinone. The indoloazepinone scaffold of pityriazepin is extremely rare in nature and has only been reported once previously. Pityriazepin, like the other isolated compounds, was found to be a potent activator of the AhR-dependent reporter gene assay in recombinant cell lines derived from four different species, although significant species differences in relative potency were observed. The ability of pityriazepin to competitively bind to the AhR and directly stimulate AhR DNA binding classified it as a new naturally-occurring potent AhR agonist. M. furfur produces an expanded collection of extremely potent naturally occurring AhR agonists, which produce their biological effects in a species-specific manner.

Advances in the in Vivo Raman Spectroscopy of Malignant Skin Tumors Using Portable Instrumentation.

Raman spectroscopy has emerged as a promising tool for real-time clinical diagnosis of malignant skin tumors offering a number of potential advantages: it is non-intrusive, it requires no sample preparation, and it features high chemical specificity with minimal water interference. However, in vivo tissue evaluation and accurate histopathological classification remain a challenging task for the successful transition from laboratory prototypes to clinical devices. In the literature, there are numerous reports on the applications of Raman spectroscopy to biomedical research and cancer diagnostics. Nevertheless, cases where real-time, portable instrumentations have been employed for the in vivo evaluation of skin lesions are scarce, despite their advantages in use as medical devices in the clinical setting. This paper reviews the advances in real-time Raman spectroscopy for the in vivo characterization of common skin lesions. The translational momentum of Raman spectroscopy towards the clinical practice is revealed by (i) assembling the technical specifications of portable systems and (ii) analyzing the spectral characteristics of in vivo measurements.

Immunocryosurgery for non-superficial basal cell carcinoma: a pro-spective, open-label phase III study for tumours ≤ 2 cm in diameter.

Cryosurgery and topical imiquimod are established mono-therapies for superficial basal cell carcinoma (BCC) but are often insufficient for thicker BCCs. We present here a phase III, prospective, interventional, single-arm (cases only) study (trial registration: NCT01212562) to evaluate the feasibility and efficacy of cryosurgery (liquid nitrogen, open spray, 2 × 15 s; day 14) during 5 weeks' imiquimod ("immunocryosurgery") for primary, non-superficial BCC, ≤ 2 cm in diameter. Ninety-one consecutive patients with 134 basal cell carcinoma were evaluated. A total of 83 patients (124 tumours) started treatment, and 79 patients (119 tumours) completed at least one cycle of immunocryosurgery (feasibility: 95.2%; follow-up: 18-60 months). The efficacy after one treatment cycle was 95 ± 2% stable complete remissions (116/119 tumours cleared, 3/116 tumours relapsed: 6 treat-ment "failures"). Neither tumour size (p = 0.865) nor localization (p = 0.233) predicted outcome. Repeat immunocryosurgery controlled 5/6 treatment failures (overall efficacy: 99%). Lack of a conventionally treated control group is a limitation of this study. However, the results show a high therapeutic efficacy of immunocryosurgery in a large series of primary non-superficial BCC.

The Malassezia genus in skin and systemic diseases.

In the last 15 years, the genus Malassezia has been a topic of intense basic research on taxonomy, physiology, biochemistry, ecology, immunology, and metabolomics. Currently, the genus encompasses 14 species. The 1996 revision of the genus resulted in seven accepted taxa: M. furfur, M. pachydermatis, M. sympodialis, M. globosa, M. obtusa, M. restricta, and M. slooffiae. In the last decade, seven new taxa isolated from healthy and lesional human and animal skin have been accepted: M. dermatis, M. japonica, M. yamatoensis, M. nana, M. caprae, M. equina, and M. cuniculi. However, forthcoming multidisciplinary research is expected to show the etiopathological relationships between these new species and skin diseases. Hitherto, basic and clinical research has established etiological links between Malassezia yeasts, pityriasis versicolor, and sepsis of neonates and immunocompromised individuals. Their role in aggravating seborrheic dermatitis, dandruff, folliculitis, and onychomycosis, though often supported by histopathological evidence and favorable antifungal therapeutic outcomes, remains under investigation. A close association between skin and Malassezia IgE binding allergens in atopic eczema has been shown, while laboratory data support a role in psoriasis exacerbations. Finally, metabolomic research resulted in the proposal of a hypothesis on the contribution of Malassezia-synthesized aryl hydrocarbon receptor (AhR) ligands to basal cell carcinoma through UV radiation-induced carcinogenesis.