Report of terbinafine resistant Trichophyton spp. in Italy: Clinical presentations, molecular identification, antifungal susceptibility testing and mutations in the squalene epoxidase gene.

BACKGROUND: Numerous reports of resistance to terbinafine in Trichophyton spp. from all over the world are arousing justified attention and concern. Point mutations in the gene that encodes the squalene epoxidase (SQLE) enzyme are responsible for these therapeutic resistances. OBJECTIVES: Primary objective of the study was to describe first isolates of Trichophyton spp. resistant to terbinafine among the patients treated between September 2019 and June 2022 at the Dermatology Units of Ospedale Maggiore Policlinico and San Bortolo Hospital. Secondary objective was to study the resistance mechanism. METHODS: Patients with confirmed Trichophyton spp. infection has been treated with systemic and topical terbinafine. Patients were then re-evaluated 12 weeks after the therapy. Patients with incomplete or absent response to terbinafine underwent a new skin scraping for direct mycological examination, new identification of dermatophyte species from culture and MALDI-TOF, molecular species identification, antifungal susceptibility testing and molecular analysis of SQLE gene. RESULTS: We identified five patients without clinical response to treatment with terbinafine. The DNA sequencing of the ITS region identified one Trichophyton rubrum and four Trichophyton indotineae. The T. rubrum strain showed minimum inhibitory concentration (MIC) (90% growth inhibition) of 4 mg/L for terbinafine. The four T. indotineae strains showed a MICs range of 0.25-4 mg/L for terbinafine. The analysis of the SQLE gene in the T. rubrum strain showed a nucleotide substitution generating a missense mutation (L393F). The SQLE gene sequencing in the T. indotineae strains showed a nucleotide substitution generating a missense mutation (F397L) in two strains, a nucleotide substitution L393S in one strain and a nucleotide substitution F415C in another strain. CONCLUSIONS: We report the first cases of terbinafine-resistant Trichophyton isolates in the Italian population. Solid antifungal management programs will be needed to promote more responsible use of antimycotics and preserve their therapeutic efficacy to control antifungal resistance.

'Colouring' wipes phenomenon: a peculiar skin pigmentation induced by ascorbic acid observed during lockdown.

During the SARS-CoV-2 (COVID-19) pandemic, an unusual outbreak of yellow-brown pigmentation on the skin of children was reported. Because of the restrictions on movement promulgated during the lockdown, most consultancies were performed using teledermatology. Data concerning personal care products and application of topical substances were collected, which revealed use of the same brand of wipes for all patients. A liquid chromatography-mass spectrometry analysis was performed to compare the components of the wipes before and after the observation of the pigmentation, in order to detect the responsible substance. This analysis revealed a level about 10-fold higher than normal of ascorbic acid and its oxidation products (dehydroascorbic acid and L-threonic acid) in the wipes associated with the pigmentation. These 'colouring wipes' represent a peculiar but harmless phenomenon that highlights the importance of careful questioning about personal care products used by patients.

Hidradenitis suppurativa in a prepubertal case series: a call for specific guidelines.

Hidradenitis suppurativa (HS) is a chronic, recurrent, inflammatory disease associated with a high physical and psychological burden. It is a disorder of the infundibular segment of the pilosebaceous unit, characterized by subcutaneous nodules, abscesses, sinus tracts and scar formation on the intertriginous and apocrine-bearing areas. HS is quite rare in young and prepubertal children. It usually begins after puberty, but several reports of prepubertal HS onset have been described. These cases are strongly linked to hormonal disorders and genetic susceptibility. Specific guidelines for prepubertal patients are still lacking, so further studies are warranted to better delineate a tailored approach. This paper aims to summarize the most significant aspects, as well as the most recent information about the epidemiology, pathogenesis, clinical features, diagnosis, comorbidities and treatment of paediatric HS. In addition, we report our clinical experience in managing HS in a group of eight prepubertal patients based on systemic antibiotics (azithromycin) and zinc oral supplementation.

Pigmented fungiform lingual papillae: dermoscopic and clinical features.

The tongue is covered by fungiform, filiform and circumvallate papillae. Fungiform papillae may be mainly pigmented in dark-skinned individuals. A single-centre study aimed to examine the clinical and dermoscopic features of pigmented fungiform papulae of the tongue (PFPT) in children, and a concise review of the literature has been performed. The clinical and anamnestic data of eight children affected by PFPT visited at the Pediatric Dermatology Unit of Bologna between 2010 and 2017, and a systemic review of all studies of PFPT published on PubMed up to 31 August 2017 has been collected and analysed. The results of our data were consistent with the literature review: dark brown to black coloured pinhead papules or bumps were observed in all cases of PFPT, and three types of clinical patterns have been detected. Moreover, the dermoscopic examination showed a cobblestone-like distribution and rose petal pattern. PFPT could be associated with hyperpigmentation of other sites such as the proximal nail folds and gums, and an intrafamiliar transmission is also possible. Clinical and dermoscopic features of PFPT may help clinicians to recognize this ethnic, acquired and benign condition.

Dose tailoring of anti-tumour necrosis factor-alpha therapy delivers useful clinical efficacy in Crohn disease patients experiencing loss of response.

BACKGROUND: 'Dose tailoring' of anti-tumour necrosis factor alpha (TNF-α) therapy in Crohn disease (CD), by dose escalation, or shortening of dosing intervals, has been suggested to regain clinical response following a flare in a proportion of patients. However, reported outcome data are sparse and none exists from Australia. METHOD: In an observational multicentre, retrospective study, the impact of anti-TNF-α dose tailoring on corticosteroid use, the need for surgery and physician perception of clinical efficacy was examined in a real-world setting at six Australian adult teaching hospitals. Demographics, disease characteristics, medications, indication for and duration of dose tailoring were documented. RESULTS: Fifty-five CD patients were identified as requiring dose tailoring and secondary loss of response was the indication in 96%. Either adalimumab (64%) or infliximab (36%) was dose escalated for a median of 5 months (range 1-47), with a median of 20 months follow up (range 3-65). At 3 months, dose tailoring reduced the mean number of days on high-dose corticosteroids (45 vs 23, P = 0.01). Most (78%) patients remained resection free, and 73% of physicians reported good clinical efficacy of dose tailoring. Of those who de-escalated therapy due to induction of remission, long-term (>12 months) follow up and complete data on steroid use were available in 15/28, with 12/15 (80%) remaining steroid free at 1 year. CONCLUSION: Short-term dose tailoring regains disease response in the majority of patients with CD. Of these, most will remain free of corticosteroids at 1 year after de-escalating therapy.

Compassionate access anti-tumour necrosis factor-α therapy for ulcerative colitis in Australia: the benefits to patients.

BACKGROUND: The efficacy of infliximab has been demonstrated in patients with both acute severe and moderate-severe ulcerative colitis (UC). However, there is a need for 'real-life data' to ensure that conclusions from trial settings are applicable in usual care. We therefore examined the national experience of anti-tumour necrosis factor-α (TNF-α) therapy in UC. METHODS: Case notes review of patients with UC who had received compassionate access (CA) anti-TNF-α therapy from prospectively maintained inflammatory bowel disease databases of six Australian adult teaching hospitals. RESULTS: Patients either received drug for acute severe UC (ASUC) failing steroids (n = 29) or for medically refractory UC (MRUC) (n = 35). In ASUC, the treating physicians judged that anti-TNF-α therapy was successful in 20/29 patients (69%); in these cases, anti-TNF-α was able to be discontinued (after 1-3 infusions in 19/20 responders) as clinical remission was achieved. Consistent with this perceived benefit, only 7/29 (24%) subsequently underwent colectomy during a median follow up of 12 months (interquartile range (IQR) 5-16). Eight of the 35 patients with MRUC (23%) required colectomy during a median follow up of 28 months (IQR 11-43). The majority of these patients (20/35 or 57%) had anti-TNF-α therapy for ≥4 months, whereas, 27/29 (93%) of ASUC patients had CA for ≤3 months. CONCLUSIONS: These data show an excellent overall benefit for anti-TNF-α therapy in both ASUC and MRUC. In particular, only short-duration anti-TNF-α was required in ASUC. These real-life data thus support the clinical trial data and should lead to broader use of this therapy in UC.

Diagnosis and Management of Intravenous Drug Users' Chronic Wounds: A Case Series Collection of a Single Center.

Background: Intravenous drug users (IVDUs) represent a very small group of patients affected by chronic wounds (CWs). Objectives: To assess the risk factors for CWs in IVDUs, to improve their treatment. Methods: A retrospective review of 7 IVDUs with CWs was performed at the Dermatology of the University of Bologna. Results: A history of 10 years of the most frequent injection was heroin in the gaiter area. The most observed comorbidities were HIV, HCV and HBV. They were treated most with an alginate with silver dressing with an average follow-up of 2.8 years. Conclusions: We would like to highlight the importance of clinical history in this type of patients and that in our experience specialized skin wounds risk assessment and management could be useful, aside from traditional compression therapy.

Automated segmentation and tracking for large-scale analysis of focal adhesion dynamics.

Cell adhesion, a process mediated by the formation of discrete structures known as focal adhesions (FAs), is pivotal to many biological events including cell motility. Much is known about the molecular composition of FAs, although our knowledge of the spatio-temporal recruitment and the relative occupancy of the individual components present in the FAs is still incomplete. To fill this gap, an essential prerequisite is a highly reliable procedure for the recognition, segmentation and tracking of FAs. Although manual segmentation and tracking may provide some advantages when done by an expert, its performance is usually hampered by subjective judgement and the long time required in analysing large data sets. Here, we developed a model-based segmentation and tracking algorithm that overcomes these problems. In addition, we developed a dedicated computational approach to correct segmentation errors that may arise from the analysis of poorly defined FAs. Thus, by achieving accurate and consistent FA segmentation and tracking, our work establishes the basis for a comprehensive analysis of FA dynamics under various experimental regimes and the future development of mathematical models that simulate FA behaviour.

The isolated comet tail pseudopodium of Listeria monocytogenes: a tail of two actin filament populations, long and axial and short and random.

Listeria monocytogenes is driven through infected host cytoplasm by a comet tail of actin filaments that serves to project the bacterium out of the cell surface, in pseudopodia, to invade neighboring cells. The characteristics of pseudopodia differ according to the infected cell type. In PtK2 cells, they reach a maximum length of approximately 15 microm and can gyrate actively for several minutes before reentering the same or an adjacent cell. In contrast, the pseudopodia of the macrophage cell line DMBM5 can extend to >100 microm in length, with the bacteria at their tips moving at the same speed as when at the head of comet tails in bulk cytoplasm. We have now isolated the pseudopodia from PtK2 cells and macrophages and determined the organization of actin filaments within them. It is shown that they possess a major component of long actin filaments that are more or less splayed out in the region proximal to the bacterium and form a bundle along the remainder of the tail. This axial component of filaments is traversed by variable numbers of short, randomly arranged filaments whose number decays along the length of the pseudopodium. The tapering of the tail is attributed to a grading in length of the long, axial filaments. The exit of a comet tail from bulk cytoplasm into a pseudopodium is associated with a reduction in total F-actin, as judged by phalloidin staining, the shedding of alpha-actinin, and the accumulation of ezrin. We propose that this transition reflects the loss of a major complement of short, random filaments from the comet, and that these filaments are mainly required to maintain the bundled form of the tail when its borders are not restrained by an enveloping pseudopodium membrane. A simple model is put forward to explain the origin of the axial and randomly oriented filaments in the comet tail.

Fyn-binding protein (Fyb)/SLP-76-associated protein (SLAP), Ena/vasodilator-stimulated phosphoprotein (VASP) proteins and the Arp2/3 complex link T cell receptor (TCR) signaling to the actin cytoskeleton.

T cell receptor (TCR)-driven activation of helper T cells induces a rapid polarization of their cytoskeleton towards bound antigen presenting cells (APCs). We have identified the Fyn- and SLP-76-associated protein Fyb/SLAP as a new ligand for Ena/ vasodilator-stimulated phosphoprotein (VASP) homology 1 (EVH1) domains. Upon TCR engagement, Fyb/SLAP localizes at the interface between T cells and anti-CD3-coated beads, where Evl, a member of the Ena/VASP family, Wiskott-Aldrich syndrome protein (WASP) and the Arp2/3 complex are also found. In addition, Fyb/SLAP is restricted to lamellipodia of spreading platelets. In activated T cells, Fyb/SLAP associates with Ena/VASP family proteins and is present within biochemical complexes containing WASP, Nck, and SLP-76. Inhibition of binding between Fyb/SLAP and Ena/VASP proteins or WASP and the Arp2/3 complex impairs TCR-dependent actin rearrangement, suggesting that these interactions play a key role in linking T cell signaling to remodeling of the actin cytoskeleton.

Annexin 2 has an essential role in actin-based macropinocytic rocketing.

Annexin 2 is a Ca(2+) binding protein that binds to and aggregates secretory vesicles at physiological Ca(2+) levels [1] and that also associates Ca(2+) independently with early endosomes [2, 3]. These properties suggest roles in both exocytosis and endocytosis, but little is known of the dynamics of Annexin 2 distribution in live cells during these processes. We have used evanescent field microscopy to image Annexin 2-GFP in live, secreting rat basophilic leukemia cells and in cells performing pinocytosis. Although we found no evidence of Annexin 2 involvement in exocytosis, we observed an enrichment of Annexin 2-GFP in actin tails propeling macropinosomes. The association of Annexin 2-GFP with rocketing macropinosomes was specific because Annexin 2-GFP was absent from the actin tails of rocketing Listeria. This finding suggests that the association of Annexin 2 with macropinocytic rockets requires native pinosomal membrane. Annexin 2 is necessary for the formation of macropinocytic rockets since overexpression of a dominant-negative Annexin 2 construct abolished the formation of these structures. The same construct did not prevent the movement of Listeria in infected cells. These results show that recruitment of Annexin 2 to nascent macropinosome membranes 16656is an essential prerequisite for actin polymerization-dependent vesicle locomotion.

The Arp2/3 complex is essential for the actin-based motility of Listeria monocytogenes.

Actin polymerisation is thought to drive the movement of eukaryotic cells and some intracellular pathogens such as Listeria monocytogenes. The Listeria surface protein ActA synergises with recruited host proteins to induce actin polymerisation, propelling the bacterium through the host cytoplasm [1]. The Arp2/3 complex is one recruited host factor [2] [3]; it is also believed to regulate actin dynamics in lamellipodia [4] [5]. The Arp2/3 complex promotes actin filament nucleation in vitro, which is further enhanced by ActA [6] [7]. The Arp2/3 complex also interacts with members of the Wiskott-Aldrich syndrome protein (WASP) [8] family - Scar1 [9] [10] and WASP itself [11]. We interfered with the targeting of the Arp2/3 complex to Listeria by using carboxy-terminal fragments of Scar1 that bind the Arp2/3 complex [11]. These fragments completely blocked actin tail formation and motility of Listeria, both in mouse brain extract and in Ptk2 cells overexpressing Scar1 constructs. In both systems, Listeria could initiate actin cloud formation, but tail formation was blocked. Full motility in vitro was restored by adding purified Arp2/3 complex. We conclude that the Arp2/3 complex is a host-cell factor essential for the actin-based motility of L. monocytogenes, suggesting that it plays a pivotal role in regulating the actin cytoskeleton.

Infliximab vs. adalimumab in Crohn's disease: results from 327 patients in an Australian and New Zealand observational cohort study.

BACKGROUND: Maintenance anti-tumour necrosis factor-α (anti-TNFα) treatment for Crohn's disease is the standard of care for patients with an inadequate response to corticosteroids and immunomodulators. AIM: To compare the efficacy and safety of infliximab and adalimumab in clinical practice and assess the value of concomitant immunomodulator therapy. METHODS: We performed an observational cohort study in consecutive patients with Crohn's disease qualifying for anti-TNFα treatment in Australia and New Zealand between 2007 and 2011. Demographic and clinical data were prospectively recorded to identify independent factors associated with induction and maintenance of response to infliximab or adalimumab, or to either anti-TNFα therapy. RESULTS: Three hundred and twenty-seven patients (183 infliximab, 144 adalimumab) successfully applied for treatment. Eighty-nine percent responded in all groups and median maintenance of response was similar for the two agents. Concomitant immunomodulator with infliximab, but not adalimumab, demonstrated a significantly longer response overall (P = 0.002), and significantly fewer disease and treatment-related complications (P = 0.017). Corticosteroids at baseline, and/or in the preceding 12 months, were associated with a 9-13 times greater risk of disease flare during maintenance treatment as compared to no corticosteroids (P < 0.0001). Maintenance of response was similar in the anti-TNF naïve and anti-TNF experienced subgroups. CONCLUSIONS: In this large, real-life study, we demonstrate infliximab and adalimumab to have similar response characteristics. However, infliximab requires concomitant immunomodulator to achieve optimal maintenance of response comparable to adalimumab monotherapy. The results of this study will assist clinicians in further optimising patient care in their day-to-day clinical practice.