Moisés Batista da Silva

Isolation of Fonsecaea pedrosoi from thorns of Mimosa pudica, a probable natural source of chromoblastomycosis

We report the isolation of Fonsecaea pedrosoi from thorns of the plant Mimosa pudica L. at the place of infection identified by one of our patients. Clinical diagnosis of chromoblastomycosis was established by direct microscopic examination and cultures from the patients lesion. The same species was isolated from the patient and from the plant. Scanning electron microscopy of the surface of the thorns showed the characteristic conidial arrangement of F. pedrosoi. These data indicate that M. pudica could be a natural source of infection for the fungus F. pedrosoi.

Cutaneous diffuse chromoblastomycosis.

A 43-year-old man presented with a 2-year history of nodules on the skin, which first appeared on the left leg and then disseminated to other regions of his skin. On examination, he had widespread nodular lesions, which were hard, elongated, violaceous, with a rough surface and well-delineated borders. In some places, such as his left buttock, the lesions became confluent, forming plaques where normal skin almost disappeared (figure, A). As well as the lesions on his limbs, the patient had lesions on his genital area, including the pubic region and the scrotum. In addition, infiltrated plaques with exulceration were found in both ear lobules. Chromoblastomycosis was confirmed by the presence of round, brown, thick-walled sclerotic bodies, which are pathognomonic for this disease (figure, B). The culture grew black filamentous fungi with a velvet surface (figure, C), and microscopic examination demonstrated cylindrical, terminal conidiophores, slackly branched, originating 3·0 1·5 m subhyaline conidia, characteristic of Fonsecaea pedrosoi (figure, D). In 1975, 25 years before his first visit to us, the patient was diagnosed with lepromatous leprosy and sent to a leprosy colony, 150 km from Belem, the state capital of Para, in the northeast of Brazil. He was treated with dapsone for many years. In 1996, he was diagnosed again with lepromatous leprosy, and he received the WHO-recommended multidrug therapy (dapsone, rifampicin, and clofazimine) for 24 months. Chromoblastomycosis is a subcutaneous mycosis with a worldwide distribution. The state of Para has the second highest prevalence rate, behind Madagascar. Usually, patients have nodular verrucous—slow growing lesions—which can appear on one of the legs and spread to neighbouring tissues through the lymphatic vessels. In this case, the lesions disseminated very rapidly, in less then 2 years, with no treatment. 5 years after the appearance of the first signs of chromoblastomycosis, the patient died of unknown causes. Cutaneous diffuse chromoblastomycosis

Cutaneous localized annular chromoblastomycosis

Chromoblastomycosis (CBM) is a difficult‐to‐treat dermal mycosis characterized by the presence of round, pigmented, sclerotic bodies formed by black fungi found in polymorphic lesions. According to the morphology of a lesion, different clinical types of the disease have been described. We present three patients who each developed a single, 10‐cm diameter, 8 to 15‐year‐old, well‐circumscribed, slow‐growing, annular, papulosquamous or papulosquamous‐verrucous lesion, with no regression despite the use of topical antifungals. Skin scrapings and biopsies confirmed CBM and microculture defined the agent as Fonsecaea pedrosoi. The patients were treated with 200 mg/day of itraconazole for 6–9 months and were discharged after complete regression of the lesions. All were examined after the first and second year of the end of treatment and there were no signs of recurrence. A new clinical type of CBM is described, and itraconazole appears to be effective and safe in curing these patients after no more than 9 months of therapy.

What do we actually know about leprosy worldwide

778 www.thelancet.com/infection Vol 16 July 2016 with leprosy varies from 1·2% to 39·8% (or why grade 2 disability ranges from 0·0% to 28·0%) in different, but all equally poor, countries? The answers will only be possible when we understand that absence of diagnosis of leprosy is not the same as the absence of leprosy. The elimination target has become the mantra everywhere, but it is now meaningless. Although the zero-transmission strategy is highly desirable, comprehension and acknowledgment of the real worldwide leprosy situation is imperative fi rst.

Molecular Epidemiology of Agents of Human Chromoblastomycosis in Brazil with the Description of Two Novel Species

The human mutilating disease chromoblastomycosis is caused by melanized members of the order Chaetothyriales. To assess population diversity among 123 clinical strains of agents of the disease in Brazil we applied sequencing of the rDNA internal transcribed spacer region, and partial cell division cycle and β-tubulin genes. Strains studied were limited to three clusters divided over the single family Herpotrichiellaceae known to comprise agents of the disease. A Fonsecaea cluster contained the most important agents, among which F. pedrosoi was prevalent with 80% of the total set of strains, followed by 13% for F. monophora, 3% for F. nubica, and a single isolate of F. pugnacius. Additional agents, among which two novel species, were located among members of the genus Rhinocladiella and Cyphellophora, with frequencies of 3% and 1%, respectively.

TGF-β plasma levels in chromoblastomycosis patients during itraconazole treatment

BACKGROUND Chromoblastomycosis (CBM) is a dermal mycosis. The disease evolves to a chronic state, presenting a suppurative granulomatous dermatitis, combined with variable dermal fibrosis. Pathogenesis of the inflammation and tissue repair in CBM are poorly understood. AIM To quantify Transforming Growth Factor-beta (TGF-beta) plasma levels of CBM patients during itraconazole (ITZ) treatment. METHODS Blood plasma of 12 CBM patients was subjected to TGF-beta titration with ELISA at 0, 3, 6 and 12months of 200mg per day of ITZ therapy, and correlated with the clinical aspects. Plasma of 12 healthy individuals were used for control. RESULTS CBM patients present high plasma levels of TGF-beta (7.016+/-1988pg/ml), decreasing after 03months (4.625+/-645pg/ml) of ITZ treatment, which correlates with a rapid clinical improvement. However, after 6 (6.566+/-777pg/ml) and 12months (6.908+/-776) of treatment, TGF-beta levels increase to almost the same levels observed before treatment, which is related to a slow clinical improvement, fungal persistence on the lesion, and fibrotic scars. CONCLUSION TGF-beta plasma levels are high in CBM patients. Fungal destruction by ITZ correlates with TGF-beta downregulation, but tissue remodeling and fungal persistence probably raises its levels again, interfering with cellular immune responses.

High Anti–Phenolic Glycolipid-I IgM Titers and Hidden Leprosy Cases, Amazon Region

To the Editor: Leprosy remains a serious public health issue. Although the World Health Organization elimination target was achieved in 2000, with a prevalence of <1 case/10,000 persons, despite progress since introduction of multidrug therapy (1), large pockets of poverty remain in which the disease is hyperendemic and underdiagnosed. In fact, in highly disease-endemic areas, the prevalence of previously undiagnosed leprosy cases in the general population has been reported to be 6× higher than the registered prevalence (2). Most leprosy patients are in India and Brazil. In Brazil, new cases are concentrated in the Northeast, Midwest, and Amazon regions (from state capitals to the inner counties). Access to the health system is poor in these regions because of severe inequalities in the public health system of Brazil (3), A total of 34,894 new cases were registered in Brazil during 2010 (4), corresponding to an incidence rate of 18.22 cases per 100,000 population. Para State accounted for 10.2% of cases (3,562 cases), an incidence rate of 46.93 per 100,000 population. When only children <15 years of age were considered, Para registered 389 new cases of leprosy in 2010, representing 10.9% of all cases, an incidence rate of 16.52 per 100,000 population. In Oriximina, a county with 62,794 inhabitants in northwestern Para, ≈800 km from Belem, Para’s capital, a mean of 13.8 cases per year were registered for the past 5 years. In 2010, in Oriximina, we collected plasma samples from 138 students 8–18 years of age, from 35 leprosy patients who received a diagnosis during 2004–2009, and from 126 contacts of these patients (Federal University of Para Research Ethics Committee protocol no. 197/07). We tested all of these samples for anti–phenolic glycolipid-I (PGL-I) IgM; 42% of students, 54.3% of case-patients, and 45% of case-patient contacts were seropositive. In addition to collecting samples, we clinically examined the leprosy patients and their contacts, among whom we identified 3 new leprosy cases. We did not examine students at that time. Contacts were persons from the same household or neighborhood whom the index case-patient described as a person with whom he or she had a close relationship. Leprosy cases were diagnosed in the field on the basis of clinical signs, loss of sensation on the skin lesions, and presence of enlarged nerves. For operational reasons, skin smears were not performed. All cases were diagnosed by 2 leprologists. We used the Ridley-Jopling classification, associated with the indeterminate clinical type, as defined by the Madrid classification. The ELISA cutoff for positive results was arbitrarily established as an optical density of 0.295 based on the average plus 3× the SD of the test results from 14 healthy persons from the Amazon region (5). Because studies of the seroprevalence among contacts have reported a proportion of seropositive persons ranging from ≈1.9% to 18.4% (6), we returned to Oriximina 16 months after the first visit. We examined 2 groups of students and their contacts; 1 group was positive for anti–PGL-I, and the other group was negative for anti–PGL-I. We visited 44 households in 1 week. From the 35 leprosy patients encountered during the first visit, we selected 25 households to survey (14 with an anti–PGL-I–positive contact in the household and 11 without), and among students with results of anti–PGL-I serology, we selected 19 households (11 positive with an anti–PGL-I–positive contact in the household and 8 without). During our visits to all of these households, we examined 222 persons (Table). Table New leprosy cases detected among selected households, Oriximina, Para State, Brazilian Amazon, 2010 When we arrived in Oriximina, only 2 cases had been registered in the national notifiable diseases information system. By using our approach, 23 new cases were found after we investigated households that had a person positive for anti–PGL-I (15 multibacillary, 8 paucibacillary); we found only 7 new cases in households where residents were negative for anti–PGL-I (4 multibacillary, 3 paucibacillary) (Table). For comparison, during the last traditional leprosy campaign in Oriximina in 2008, eight new cases were detected. Furthermore, by using our strategy, the local public health service detected 9 additional new cases during the 4 months after our departure from Oriximina. These data emphasize that contact examination is crucial for identifying new cases (7) and that such investigation must be conducted periodically. Our data also indicate that subclinical infections are highly prevalent among public school students in the Amazon region and that identifying students with positive anti–PGL-I test results can lead to discovery of new leprosy cases among students’ household contacts.

Field-friendly serological tests for determination of M. leprae -specific antibodies

Early detection of leprosy is key to reduce the ongoing transmission. Antibodies directed against M. leprae PGL-I represent a useful biomarker for detecting multibacillary (MB) patients. Since efficient leprosy diagnosis requires field-friendly test conditions, we evaluated two rapid lateral flow assays (LFA) for detection of Mycobacterium leprae-specific antibodies: the visual immunogold OnSite Leprosy Ab Rapid test [Gold-LFA] and the quantitative, luminescent up-converting phosphor anti-PGL-I test [UCP-LFA]. Test performance was assessed in independent cohorts originating from three endemic areas. In the Philippine cohort comprising patients with high bacillary indices (BI; average:4,9), 94%(n = 161) of MB patients were identified by UCP-LFA and 78%(n = 133) by Gold-LFA. In the Bangladeshi cohort, including mainly MB patients with low BI (average:1), 41%(n = 14) and 44%(n = 15) were detected by UCP-LFA and Gold-LFA, respectively. In the third cohort of schoolchildren from a leprosy hyperendemic region in Brazil, both tests detected 28%(n = 17) seropositivity. Both rapid tests corresponded well with BI(p < 0.0001), with a fairly higher sensitivity obtained with the UCP-LFA assay. However, due to the spectral character of leprosy, additional, cellular biomarkers are required to detect patients with low BIs. Therefore, the UCP-LFA platform, which allows multiplexing with differential biomarkers, offers more cutting-edge potential for diagnosis across the whole leprosy spectrum.

Enzymatic isolation of Lacazia loboi cells from skin lesions of lobomycosis

Lacazia loboi is the etiologic agent of Jorge Lobos disease, a cutaneous and subcutaneous mycosis endemic to Latin America tropical regions and characterized by chronic nodular or keloidal lesions which develop after traumatic events. A new method for the extraction of L. loboi yeast cells from biopsies of lobomycosis skin lesions is presented. The method is based on the proteolytic action of the enzyme dispase which is known for its action against fibronectin and collagen type IV. Fungal identification was based on histological examination of the biological material and molecular analysis based on 18S ribosomal sequences. Observations under optic and fluorescence microscopy proved the efficacy of enzymatic isolation of the lobomycosis etiologic agent, as well as identifying the organisms main parasitic characteristics. Molecular phylogenetic analysis corroborated the histological examination and indicated L. loboi relationship with other members of the Onygenales. Use of dispase proved to be ideal for the isolation of L. loboi from human biopsies, shows promise as an important tool for improving biological studies of this peculiar fungus.

Effects of immunomodulatory drugs on TNF-α and IL-12 production by purified epidermal langerhans cells and peritoneal macrophages

BackgroundLangerhans cells constitute a special subset of immature dendritic cells localized in the epidermis that play a key role in the skins immune response. The production of cytokines is a key event in both the initiation and the regulation of immune responses, and different drugs can be used to remove or modify their production by DC and, therefore, alter immune responses in a broad spectrum of diseases, mainly in human inflammatory and autoimmune diseases. In the present study, we examined the effects of prednisone, thalidomide, cyclosporine A, and amitriptyline, drugs used in a variety of clinical conditions, on the production of TNF-α, IL-10, and IL-12 by purified epidermal Langerhans cells and peritoneal macrophages in BALB/c mice.FindingsAll drugs inhibited TNF-α production by Langerhans cells after 36 hours of treatment at two different concentrations, while prednisone and thalidomide decreased IL-12 secretion significantly, amitriptyline caused a less pronounced reduction and cyclosporine A had no effect. Additionally, TNF-α and IL-12 production by macrophages decreased, but IL-10 levels were unchanged after all treatments.ConclusionsOur results demonstrate that these drugs modulate the immune response by regulating pro-inflammatory cytokine production by purified epidermal Langerhans cells and peritoneal macrophages, indicating that these cells are important targets for immunosuppression in various clinical settings.