The Effect of Geoclimatic Factors on the Distribution of Paracoccidioidomycosis in Mato Grosso do Sul, Brazil.

The incidence of paracoccidioidomycosis (PCM) varies in Latin America, and it is influenced by environmental factors. This study evaluated the distribution of PCM acute/subacute form (AF) cases and their correlation with geoclimatic factors in the Mato Grosso do Sul (MS) state. The study included 81 patients diagnosed with the PCM/AF at the University Hospital of the Federal University of Mato Grosso do Sul between January 1980 and February 2022. Geographic coordinates, health microregion of patient's residence, compensated average temperature, relative air humidity (RH), El Niño Southern Oscillation (ENSO), and average global temperature were analyzed. The highest incidence was observed in the Aquidauana (7/100,000 inhabitants), while Campo Grande, the state's capital, had the highest number (n = 34; 42.4%) and density (4.4 cases/km2) of cases. The number of cases increased during extended periods of the El Niño phenomenon. A positive correlation was found between higher RH and PCM/AF cases. Most PCM/AF cases were found in areas with loamy soils and RH ranging from 60.8 to 73.6%. In MS, the health microregions of PCM/AF patients are characterized by deforestation for agricultural and pasture use, coupled with loamy soils and specific climatic phenomena leading to higher soil humidity.

Genetic diversity of Mycobacterium leprae in the state of São Paulo, an area of low-leprosy incidence in Brazil.

BACKGROUND: Brazil has the second largest number of leprosy cases worldwide, and the state of São Paulo has been considered non-endemic since 2006. METHODS: We analyzed 16 variable number tandem repeats loci and three single nucleotide polymorphisms loci of Mycobacterium leprae (M. leprae) in 125 clinical isolates from patients in different municipalities in the state. RESULTS: The clustering pattern of M. leprae indicated that the transmission of leprosy persisted in the state and included scenarios of intra-extra-familial transmission in areas with low endemicity. CONCLUSIONS: A significantly active circulation of M. leprae was observed. Therefore, surveillance and control measures must be implemented.

Genetic diversity of Mycobacterium leprae in the state of São Paulo, an area of low-leprosy incidence in Brazil

ABSTRACT Background: Brazil has the second largest number of leprosy cases worldwide, and the state of São Paulo has been considered non-endemic since 2006. Methods: We analyzed 16 variable number tandem repeats loci and three single nucleotide polymorphisms loci of Mycobacterium leprae (M. leprae) in 125 clinical isolates from patients in different municipalities in the state. Results: The clustering pattern of M. leprae indicated that the transmission of leprosy persisted in the state and included scenarios of intra-extra-familial transmission in areas with low endemicity. Conclusions: A significantly active circulation of M. leprae was observed. Therefore, surveillance and control measures must be implemented.

Social school contacts of multibacillary leprosy cases in children living in the hyperendemic region of the Midwest of Brazil

Abstract Objective: To analyze the prevalence of Mycobacterium leprae detection and the associated factors among social contacts in the school environment of multibacillary cases living in a hyperendemic municipality of the state of Mato Grosso. Methods: Cross-sectional study with 236 social contacts of multibacillary leprosy from public schools and residents in Cuiabá (Mato Grosso) in 2018. The sources of information were interviews and nasal swab tests for molecular analysis by polymerase chain reaction - PCR. For the prevalence ratio estimates, crude and adjusted analyses were performed using robust Poisson regression and their respective confidence intervals (95% CI). The ArcGIS 9.1 software was used for the geographic distribution analyses. Results: The prevalence of detection of M. leprae in social contacts was 14%. A total of 63.6% of the schools surveyed had 5.1% to 50% of the social contacts of leprosy with positive PCR. The analysis of the geographic distribution in the neighborhoods showed a high prevalence of infection, being higher than 50% in some localities. The highest proportion of positive results occurred in the northern region of the city and from a precarious socioeconomic class. Conclusion: The results showed a high prevalence of detection of M. leprae among social contacts in areas with poor socioeconomic conditions. In these regions, there is a greater risk of

Social school contacts of multibacillary leprosy cases in children living in the hyperendemic region of the Midwest of Brazil.

OBJECTIVE: To analyze the prevalence of Mycobacterium leprae detection and the associated factors among social contacts in the school environment of multibacillary cases living in a hyperendemic municipality of the state of Mato Grosso. METHODS: Cross-sectional study with 236 social contacts of multibacillary leprosy from public schools and residents in Cuiabá (Mato Grosso) in 2018. The sources of information were interviews and nasal swab tests for molecular analysis by polymerase chain reaction - PCR. For the prevalence ratio estimates, crude and adjusted analyses were performed using robust Poisson regression and their respective confidence intervals (95% CI). The ArcGIS 9.1 software was used for the geographic distribution analyses. RESULTS: The prevalence of detection of M. leprae in social contacts was 14%. A total of 63.6% of the schools surveyed had 5.1% to 50% of the social contacts of leprosy with positive PCR. The analysis of the geographic distribution in the neighborhoods showed a high prevalence of infection, being higher than 50% in some localities. The highest proportion of positive results occurred in the northern region of the city and from a precarious socioeconomic class. CONCLUSION: The results showed a high prevalence of detection of M. leprae among social contacts in areas with poor socioeconomic conditions. In these regions, there is a greater risk of infection and of getting sick.

Virulence Factors and Antifungal Susceptibility in Candida Species Isolated from Dermatomycosis Patients.

Dermatomycoses caused by Candida spp. are increasingly common, however there are few reports in the literature regarding their epidemiology, pathogenesis and antifungal susceptibility of these fungal pathogens. This study aimed to identify, characterize virulence and determine the antifungal susceptibility of Candida species isolated from patients with dermatomycoses. We studied a total of 45 yeast samples isolated from dermal scrapings from patients treated at a public hospital, Bauru, São Paulo, Brazil. After identification, the samples were analyzed for protease activity, phospholipase, biofilm formation and antifungal susceptibility. The most common species was Candida albicans (40%), followed by C. krusei (22.22%), C. parapsilosis complex (22.22%) and C. famata (6.67%). Regarding virulence mechanisms, 57.78% and 28.89% of the samples showed enzymatic activity for protease and phospholipase, respectively. A total of 13.33% of the samples showed secretion of both enzymes. All isolates were biofilm producers, and a higher production was observed in C. tropicalis isolates. Regarding the antifungal susceptibility test, 97.78% of the samples had the highest levels of the minimal inhibitory concentration (MIC) for terbinafine, and one C. krusei sample showed intermediate susceptibility to fluconazole. The correct identification of dermatomycosis-related microorganisms, as well as a better knowledge of their pathogenicity and susceptibility against the antifungals used in the clinic, is extremely important for an efficient preventive and therapeutic action.

Perfil sociodemográfico, clínico e geoespacial de casos novos de hanseníase diagnosticados no Instituto Lauro de Souza Lima, Bauru, São Paulo, entre 2015 e 2019 / Sociodemographic, clinical and geospatial profile of new leprosy cases diagnosed at Institute Lauro de Souza Lima, Bauru, São Paulo, between 2015 and 2019

Embora o Brasil venha apresentando uma redução considerável no número de casos novos de hanseníase, alguns estudos têm demonstrado transmissão ativa mesmo em áreas não endêmicas, como o estado de São Paulo. Diante disso, investigamos o perfil sociodemográfico, clínico e geoespacial dos casos novos de hanseníase diagnosticados entre 2015 e 2019 no Instituto Lauro de Souza Lima (ILSL), um centro de referência localizado no município de Bauru, interior do estado de São Paulo. Foram diagnosticados 177 novos casos de hanseníase nesse período, sendo 61,6% dos pacientes naturais do estado. A maioria dos indivíduos era do sexo masculino (59,9%) e a faixa etária mais prevalente foi de 60 a 69 anos; 79,1% se autodeclaravam brancos e 65,6% possuíam pouca ou nenhuma escolaridade. A forma clínica dimorfa foi a mais frequente (42,4%), a baciloscopia foi positiva em 38,4% dos pacientes (49,0% entre o sexo masculino e 22,5% entre o sexo feminino) e 49,0% dos pacientes possuíam incapacidades no diagnóstico. O georreferenciamento, realizado para os casos oriundos do município de Bauru (n = 31), revelou que a maioria dos pacientes residia em regiões com elevado nível de vulnerabilidade social. O perfil dos pacientes atendidos no ILSL apontou para predominância de homens adultos ou idosos com baixa escolaridade, multibacilares, apresentando incapacidades físicas e longo tempo de sintomas. Em conjunto, nossos dados sugerem atraso no diagnóstico que pode contribuir para a manutenção da transmissão da hanseníase mesmo numa região não endêmica.(AU)

Although Brazil has shown a considerable reduction in the number of new cases of leprosy, some studies have shown active transmission even in non-endemic areas, such as São Paulo state. Considering this, we investigated the sociodemographic, clinical and geospatial profile of new cases of leprosy diagnosed between 2015 and 2019 at the Lauro de Souza Lima Institute (ILSL) a reference center localized in Bauru, a municipality in the interior of São Paulo state. A total of 177 new cases of leprosy were diagnosed in this period, with 61.6% of the patients born in the state. The majority of the individuals was male (59.9%), the most prevalent age group was 60 to 69 years old, 79.1% declared themselves white color/race and 65.6% had little or no schooling. The borderline form of leprosy was the most frequent (42.4%), slit skin smear was positive in 38.4% of patients (49.0% among men and 22.5% among women) and 49.0% had disabilities at diagnosis. Georeferencing, performed for cases from the municipality of Bauru (n = 31), revealed that most patients lived in regions with a high level of social vulnerability. The profile of leprosy new cases found at ILSL pointed to a predominance of adult or elderly men with low education, multibacillary, presenting physical disabilities and a long time of symptoms. Altogether, our data suggests a delay in diagnosis that may contribute to the maintenance of leprosy transmission even in a non-endemic region.(AU)

Virulence factors and antifungal susceptibility in candida species isolated from dermatomycosis patients

Dermatomycoses caused by Candida spp. are increasingly common, however there are few reports in the literature regarding their epidemiology, pathogenesis and antifungal susceptibility of these fungal pathogens. This study aimed to identify, characterize virulence and determine the antifungal susceptibility of Candida species isolated from patients with dermatomycoses. We studied a total of 45 yeast samples isolated from dermal scrapings from patients treated at a public hospital, Bauru, São Paulo, Brazil. After identification, the samples were analyzed for protease activity, phospholipase, biofilm formation and antifungal susceptibility. The most common species was Candida albicans (40%), followed by C. krusei (22.22%), C. parapsilosis complex (22.22%) and C. famata (6.67%). Regarding virulence mechanisms, 57.78% and 28.89% of the samples showed enzymatic activity for protease and phospholipase, respectively. A total of 13.33% of the samples showed secretion of both enzymes. All isolates were biofilm producers, and a higher production was observed in C. tropicalis isolates. Regarding the antifungal susceptibility test, 97.78% of the samples had the highest levels of the minimal inhibitory concentration (MIC) for terbinafine, and one C. krusei sample showed intermediate susceptibility to fluconazole. The correct identification of dermatomycosis-related microorganisms, as well as a better knowledge of their pathogenicity and susceptibility against the antifungals used in the clinic, is extremely important for an efficient preventive and therapeutic action(AU).

Phylogenomics and antimicrobial resistance of the leprosy bacillus Mycobacterium leprae

Leprosy is a chronic human disease caused by the yet-uncultured pathogen Mycobacterium leprae. Although readily curable with multidrug therapy (MDT), over 200,000 new cases are still reported annually. Here, we obtain M. leprae genome sequences from DNA extracted directly from patients' skin biopsies using a customized protocol. Comparative and phylogenetic analysis of 154 genomes from 25 countries provides insight into evolution and antimicrobial resistance, uncovering lineages and phylogeographic trends, with the most ancestral strains linked to the Far East. In addition to known MDT-resistance mutations, we detect other mutations associated with antibiotic resistance, and retrace a potential stepwise emergence of extensive drug resistance in the pre-MDT era. Some of the previously undescribed mutations occur in genes that are apparently subject to positive selection, and two of these (ribD, fadD9) are restricted to drug-resistant strains. Finally, nonsense mutations in the nth excision repair gene are associated with greater sequence diversity and drug resistance.

Identificação, fatores de virulência e susceptibilidade a drogas em espécies de candida isoladas de pacientes atendidos no Instituto Lauro de Souza Lima / Identification, virulence factors and susceptibility to drugs in candida species isolated from patients attending the Instituto Lauro de Souza Lima

Dentre as dermatomicoses, as micoses causadas por leveduras do gênero Candida são cada vez mais frequentes, sendo a espécie Candida albicans de maior prevalência, porém as espécies não-albicans vem apresentando um aumento gradativo nas dermatomicoses. O aumento destas espécies vem sendo preocupante nos ambientes hospitalares assim a identificação destes microrganismo é de extrema importância para uma ação preventiva e terapêutica eficiente. Objetivo: Realizar a identificação das espécies através de técnicas bioquímicas, identificar mecanismos de virulências e caracterizar, entre as espécies sua susceptibilidade aos diferentes antifúngicos. Material e métodos: A identificação das espécies de Candida das amostras de raspados dérmicos foi realizada pelo cultivo em CHROMagar* e as técnicas de fermentação e assimilação de açucares. Os mecanismos de virulência foram avaliados para fosfolipases, proteinases e biofilme. O padrão de sensibilidade das amostras foi avaliado pela utilização da metodologia de microdiluição. Resultado: O CHROMagar* foi utilizado como identificação presuntiva para avaliar pureza das colônias, a identificação através da assimilação e fermentação de açucares caracterizou 40% de C. albicans, 22,22% de C. krusei, 22,22% de C. parapsilosis, 6,67% de C. famata, 4,44% de C. tropicalis, 2,22% de C. lusitaniae e 2,22% de C. zeylanoides. Para os mecanismos de virulência 57,78% das amostras isoladas apresentaram produção de proteinase e 28,89% apresentaram produção para fosfolipase e um total de 13,33% do total das amostras isoladas foram produtoras de ambas enzimas. Para produção de biofilme todas foram produtoras, sendo as C. tropicalis as maiores produtoras. para os testes de susceptibilidade aos antifúngicos, foram encontrados altos valores de MIC’s para a terbinafina em 97,78% das amostras, e uma amostra de C. krusei apresentou susceptibilidade intermediaria para fluconazol...

Among the dermatomycoses, mycoses caused by yeasts of the genus Candida are more frequent, with the Candida albicans species being more prevalent, but the non-albicans species have been presenting a gradual increase in the dermatomycoses. The increase of these species has been worrying in the hospital environments so the identification of these microorganisms is of extreme importance for an efficient preventive and therapeutic action. Objective: Identify the species through biochemical techniques, identify mechanisms of virulence and characterize, among the species, their susceptibility to different antifungal agents. Material and methods: Identification of the Candida species of the dermal scrap samples was performed by the cultivation in CHROMagar * and the techniques of fermentation and assimilation of sugars. Virulence mechanisms were evaluated for phospholipases, proteinases and biofilms. The sensitivity standard of the samples was evaluated using the microdilution methodology. Results: CHROMagar* was used as presumptive identification to evaluate the purity of the colonies. Identification through assimilation and fermentation of sugars characterized 40% of C. albicans, 22.22% of C. krusei, 22.22% of C. parapsilosis, 6.67% of C. famata, 4.44% of C. tropicalis, 2.22% of C. lusitaniae and 2.22% of C. zeylanoides. For the virulence mechanisms, 57.78% of the isolated samples presented proteinase production and 28.89% presented phospholipase production and a total of 13.33% of the total of the isolated samples were producing both enzymes. For biofilm production, all were producers, being C. tropicalis the largest producers. for antifungal susceptibility tests, high MIC values for terbinafine were found in 97.78% of the samples, and a sample of C. krusei showed intermediate susceptibility to fluconazole...

Tuberculosis associated factors caused by Mycobacterium tuberculosis of the RDRio genotype

BACKGROUND Tuberculosis (TB) continues to be a disease that affects many countries around the world, including Brazil. Recently, a subtype of Latin American-Mediterranean family strain was identified and characterised by RDRio. The strain has been associated with different characteristics of the disease. OBJECTIVES In the present study we investigated the association of epidemiological, clinical, radiological and bacteriological variables with pulmonary tuberculosis caused by RDRioMycobacterium tuberculosis strain in large regions of São Paulo. METHODS We conducted a cross-sectional study in 530 patients with pulmonary tuberculosis, diagnosed using sputum culture, from two regions of the São Paulo state in Brazil. The samples were brought to São Paulo reference laboratories for epidemiological, clinical, radiological and bacteriological analyses, and the data were obtained from a TB notification system. RDRio genotyping and Spoligotyping of the samples were performed. For the analysis of the categorical variables we used the chi-square test or the Fisher’s exact test, and for the continuous variables, the Mann-Whitney test. In addition, a logistic regression was used for multivariate analysis. Differences with p < 0.05 were considered significant. FINDINGS The RDRio deletion was identified in 152 (28.7%) samples. In the univariate analysis, both the age groups above 25 years and alcohol consumption were associated with the RDRio deletion. The multivariate analysis confirmed the association of the RDRio deletion with the age groups: 25-35 years old [OR: 2.28 (1.02-5.07; p = 0.04)] and 36-60 years old (OR: 2.36 (1.11-5.05); p = 0.03], and also with alcohol consumption [OR: 1.63 (1.05-2.54); p = 0,03]. MAIN CONCLUSIONS In this study, we identified new factors associated with the M. tuberculosis of the RDRio deletion strains infection.

Tuberculosis associated factors caused by Mycobacterium tuberculosis of the RDRio genotype.

BACKGROUND: Tuberculosis (TB) continues to be a disease that affects many countries around the world, including Brazil. Recently, a subtype of Latin American-Mediterranean family strain was identified and characterised by RDRio. The strain has been associated with different characteristics of the disease. OBJECTIVES: In the present study we investigated the association of epidemiological, clinical, radiological and bacteriological variables with pulmonary tuberculosis caused by RDRioMycobacterium tuberculosis strain in large regions of São Paulo. METHODS: We conducted a cross-sectional study in 530 patients with pulmonary tuberculosis, diagnosed using sputum culture, from two regions of the São Paulo state in Brazil. The samples were brought to São Paulo reference laboratories for epidemiological, clinical, radiological and bacteriological analyses, and the data were obtained from a TB notification system. RDRio genotyping and Spoligotyping of the samples were performed. For the analysis of the categorical variables we used the chi-square test or the Fisher's exact test, and for the continuous variables, the Mann-Whitney test. In addition, a logistic regression was used for multivariate analysis. Differences with p < 0.05 were considered significant. FINDINGS THE RDRIO: deletion was identified in 152 (28.7%) samples. In the univariate analysis, both the age groups above 25 years and alcohol consumption were associated with the RDRio deletion. The multivariate analysis confirmed the association of the RDRio deletion with the age groups: 25-35 years old [OR: 2.28 (1.02-5.07; p = 0.04)] and 36-60 years old (OR: 2.36 (1.11-5.05); p = 0.03], and also with alcohol consumption [OR: 1.63 (1.05-2.54); p = 0,03]. MAIN CONCLUSIONS: In this study, we identified new factors associated with the M. tuberculosis of the RDRio deletion strains infection.

Tuberculosis associated factors caused by Mycobacterium tuberculosis of the RDrio genotype

BACKGROUND: Tuberculosis (TB) continues to be a disease that affects many countries around the world, including Brazil. Recently, a subtype of Latin American-Mediterranean family strain was identified and characterised by RDRio. The strain has been associated with different characteristics of the disease. OBJECTIVES: In the present study we investigated the association of epidemiological, clinical, radiological and bacteriological variables with pulmonary tuberculosis caused by RDRioMycobacterium tuberculosis strain in large regions of São Paulo. METHODS: We conducted a cross-sectional study in 530 patients with pulmonary tuberculosis, diagnosed using sputum culture, from two regions of the São Paulo state in Brazil. The samples were brought to São Paulo reference laboratories for epidemiological, clinical, radiological and bacteriological analyses, and the data were obtained from a TB notification system. RDRio genotyping and Spoligotyping of the samples were performed. For the analysis of the categorical variables we used the chi-square test or the Fisher's exact test, and for the continuous variables, the Mann-Whitney test. In addition, a logistic regression was used for multivariate analysis. Differences with p < 0.05 were considered significant. FINDINGS THE RDRIO: deletion was identified in 152 (28.7%) samples. In the univariate analysis, both the age groups above 25 years and alcohol consumption were associated with the RDRio deletion. The multivariate analysis confirmed the association of the RDRio deletion with the age groups: 25-35 years old [OR: 2.28 (1.02-5.07; p = 0.04)] and 36-60 years old (OR: 2.36 (1.11-5.05); p = 0.03], and also with alcohol consumption [OR: 1.63 (1.05-2.54); p = 0,03]. MAIN CONCLUSIONS: In this study, we identified new factors associated with the M. tuberculosis of the RDRio deletion strains infection.

Clinical and epidemiological profiles of individuals with drug-resistant tuberculosis.

Drug-resistant tuberculosis (TB) is a growing global threat. Approximately 450,000 people developed multidrug-resistant TB worldwide in 2012 and an estimated 170,000 people died from the disease. This paper describes the sociodemographic, clinical-epidemiological and bacteriological aspects of TB and correlates these features with the distribution of anti-TB drug resistance. Mycobacterium tuberculosis (MT) cultures and drug susceptibility testing were performed according to the BACTEC MGIT 960 method. The results demonstrated that MT strains from individuals who received treatment for TB and people who were infected with human immunodeficiency virus were more resistant to TB drugs compared to other individuals (p < 0.05). Approximately half of the individuals received supervised treatment, but most drug-resistant cases were positive for pulmonary TB and exhibited positive acid-fast bacilli smears, which are complicating factors for TB control programs. Primary healthcare is the ideal level for early disease detection, but tertiary healthcare is the most common entry point for patients into the system. These factors require special attention from healthcare managers and professionals to effectively control and monitor the spread of TB drug-resistant cases.

Clinical and epidemiological profiles of individuals with drug-resistant tuberculosis

Drug-resistant tuberculosis (TB) is a growing global threat. Approximately 450,000 people developed multidrug-resistant TB worldwide in 2012 and an estimated 170,000 people died from the disease. This paper describes the sociodemographic, clinical-epidemiological and bacteriological aspects of TB and correlates these features with the distribution of anti-TB drug resistance. Mycobacterium tuberculosis (MT) cultures and drug susceptibility testing were performed according to the BACTEC MGIT 960 method. The results demonstrated that MT strains from individuals who received treatment for TB and people who were infected with human immunodeficiency virus were more resistant to TB drugs compared to other individuals (p < 0.05). Approximately half of the individuals received supervised treatment, but most drug-resistant cases were positive for pulmonary TB and exhibited positive acid-fast bacilli smears, which are complicating factors for TB control programs. Primary healthcare is the ideal level for early disease detection, but tertiary healthcare is the most common entry point for patients into the system. These factors require special attention from healthcare managers and professionals to effectively control and monitor the spread of TB drug-resistant cases.

Analysis of immune response markers in Jorge Lobo's disease lesions suggests the occurrence of mixed T Helper responses with the dominance of regulatory T cell activity

Jorge Lobo's disease (JLD) is a chronic infection that affects the skin and subcutaneous tissues. Its etiologic agent is the fungus Lacazia loboi. Lesions are classified as localized, multifocal, or disseminated, depending on their location. Early diagnosis and the surgical removal of lesions are the best therapeutic options currently available for JLD. The few studies that evaluate the immunological response of JLD patients show a predominance of Th2 response, as well as a high frequency of TGF-ß and IL-10 positive cells in the lesions; however, the overall immunological status of the lesions in terms of their T cell phenotype has yet to be determined. Therefore, the objective of this study was to evaluate the pattern of Th1, Th2, Th17 and regulatory T cell (Treg) markers mRNA in JLD patients by means of real-time PCR. Biopsies of JLD lesions (N = 102) were classified according to their clinical and histopathological features and then analyzed using real-time PCR in order to determine the expression levels of TGF-ß1, FoxP3, CTLA4, IKZF2, IL-10, T-bet, IFN-γ, GATA3, IL-4, IL-5, IL-13, IL-33, RORC, IL-17A, IL-17F, and IL-22 and to compare these levels to those of healthy control skin (N = 12). The results showed an increased expression of FoxP3, CTLA4, TGF-ß1, IL-10, T-bet, IL-17F, and IL-17A in lesions, while GATA3 and IL-4 levels were found to be lower in diseased skin than in the control group. When the clinical forms were compared, TGF-ß1 was found to be highly expressed in patients with a single localized lesion while IL-5 and IL-17A levels were higher in patients with multiple/disseminated lesions. These results demonstrate the occurrence of mixed T helper responses and suggest the dominance of regulatory T cell activity, which could inhibit Th-dependent protective responses to intracellular fungi such as L. loboi. Therefore, Tregs may play a key role in JLD pathogenesis.

Pre-miR 146a (rs2910164 G>C) single nucleotide polymorphism is genetically and functionally associated with leprosy

Mycobacterium leprae infects macrophages and Schwann cells inducing a gene expression program to facilitate its replication and progression to disease. MicroRNAs (miRNAs) are key regulators of gene expression and could be involved during the infection. To address the genetic influence of miRNAs in leprosy, we enrolled 1,098 individuals and conducted a case-control analysis in order to study four miRNAs genes containing single nucleotide polymorphism (miRSNP). We tested miRSNP-125a (rs12975333 G>T), miRSNP-223 (rs34952329 *>T), miRSNP-196a-2 (rs11614913 C>T) and miRSNP-146a (rs2910164 G>C). Amongst them, miRSNP-146a was the unique gene associated with risk to leprosy per se (GC OR = 1.44, p = 0.04; CC OR = 2.18, p = 0.0091). We replicated this finding showing that the C-allele was over-transmitted (p = 0.003) using a transmission-disequilibrium test. A functional analysis revealed that live M. leprae (MOI 100:1) was able to induce miR-146a expression in THP-1 (p<0.05). Furthermore, pure neural leprosy biopsies expressed augmented levels of that miRNA as compared to biopsy samples from neuropathies not related with leprosy (p = 0.001). Interestingly, carriers of the risk variant (C-allele) produce higher levels of mature miR-146a in nerves (p = 0.04). From skin biopsies, although we observed augmented levels of miR-146a, we were not able to correlate it with a particular clinical form or neither host genotype. MiR-146a is known to modulate TNF levels, thus we assessed TNF expression (nerve biopsies) and released by peripheral blood mononuclear cells infected with BCG Moreau. In both cases lower TNF levels correlates with subjects carrying the risk C-allele, (p = 0.0453 and p = 0.0352; respectively), which is consistent with an immunomodulatory role of this miRNA in leprosy.

Genotyping of Mycobacterium leprae present on Ziehl-Neelsen-stained microscopic slides and in skin biopsy samples from leprosy patients in different geographic regions of Brazil

We analysed 16 variable number tandem repeats (VNTR) and three single-nucleotide polymorphisms (SNP) in Mycobacterium leprae present on 115 Ziehl-Neelsen (Z-N)-stained slides and in 51 skin biopsy samples derived from leprosy patients from Ceará (n = 23), Pernambuco (n = 41), Rio de Janeiro (n = 22) and Rondônia (RO) (n = 78). All skin biopsies yielded SNP-based genotypes, while 48 of the samples (94.1%) yielded complete VNTR genotypes. We evaluated two procedures for extracting M. leprae DNA from Z-N-stained slides: the first including Chelex and the other combining proteinase and sodium dodecyl sulfate. Of the 76 samples processed using the first procedure, 30.2% were positive for 16 or 15 VNTRs, whereas of the 39 samples processed using the second procedure, 28.2% yielded genotypes defined by at least 10 VNTRs. Combined VNTR and SNP analysis revealed large variability in genotypes, but a high prevalence of SNP genotype 4 in the Northeast Region of Brazil. Our observation of two samples from RO with an identical genotype and seven groups with similar genotypes, including four derived from residents of the same state or region, suggest a tendency to form groups according to the origin of the isolates. This study demonstrates the existence of geographically related M. leprae genotypes and that Z-N-stained slides are an alternative source for M. leprae genotyping.

Genotyping of Mycobacterium leprae present on Ziehl-Neelsen-stained microscopic slides and in skin biopsy samples from leprosy patients in different geographic regions of Brazil.

We analysed 16 variable number tandem repeats (VNTR) and three single-nucleotide polymorphisms (SNP) in Mycobacterium leprae present on 115 Ziehl-Neelsen (Z-N)-stained slides and in 51 skin biopsy samples derived from leprosy patients from Ceará (n = 23), Pernambuco (n = 41), Rio de Janeiro (n = 22) and Rondônia (RO) (n = 78). All skin biopsies yielded SNP-based genotypes, while 48 of the samples (94.1%) yielded complete VNTR genotypes. We evaluated two procedures for extracting M. leprae DNA from Z-N-stained slides: the first including Chelex and the other combining proteinase and sodium dodecyl sulfate. Of the 76 samples processed using the first procedure, 30.2% were positive for 16 or 15 VNTRs, whereas of the 39 samples processed using the second procedure, 28.2% yielded genotypes defined by at least 10 VNTRs. Combined VNTR and SNP analysis revealed large variability in genotypes, but a high prevalence of SNP genotype 4 in the Northeast Region of Brazil. Our observation of two samples from RO with an identical genotype and seven groups with similar genotypes, including four derived from residents of the same state or region, suggest a tendency to form groups according to the origin of the isolates. This study demonstrates the existence of geographically related M. leprae genotypes and that Z-N-stained slides are an alternative source for M. leprae genotyping.