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Introduction. The increased production of new drugs is parallel with the bacterial adaptation to adverse conditions. There are several factors that have been pointed out as the cause of this, such as: globalization; the abusive use of antibiotics; and the arbitrary prescription of these drugs. Therefore, the objective of the present study was to evaluate antimicrobial usage with antibiotic resistance in 2012 and 2013 at a university hospital in Southern Brazil. Material and Methods. This study was performed at the University Hospital Dr. Miguel Riet Correa Jr. in Rio Grande/RS, Brazil. The data were collected between January 2012 and December 2013, using data from the medical clinic and two intensive care units. Results. Were observed, 385 and 464 cases of bacterial infections, during the years of 2012 and 2013, respectively. Acinetobacter baumannii, Escherichia coli and Klebsiella pneumoniae were the most prevalent species. The total antibiotic consumption in the University Hospital (HU-FURG) in this period was 3.865 units/dose, with 1.995 units/dose in the year 2012 and 1.870 units/dose in the year 2013. Relationships between the rates of main resistant pathogens isolated and the annual consumption of the corresponding antibiotic (carbapenems, cephalosporins, sulfonamides and other β-lactams) were observed; however, they did not show significant positive associations in increased resistance associated with increased consumption. Conclusion. This study reveals the need for the dissemination and feedback of these data to clinicians and decision-makers at the hospital, as it may be crucial to improve policies on prescribing antibiotics and to implement effective infection control. Introducción. El aumento en la producción de nuevos medicamentos surge en paralelo con la adaptación bacteriana a las condiciones adversas. Hay varios factores que se han identificado como la causa de esta afección, como: globalización, abuso de antibióticos y prescripción arbitraria de estos medicamentos. Por lo tanto, el objetivo del presente estudio fue evaluar el uso de antimicrobianos con resistencia a los antibióticos en 2012 y 2013 en un hospital universitario en el Sur de Brasil. Material y métodos. Este estudio se realizó en el Hospital Universitario Dr. Miguel Riet Correa Jr. en Rio Grande / RS, Brasil. Los datos se recopilaron entre enero de 2012 y diciembre de 2013, en la clínica médica y dos unidades de cuidados intensivos. Resultados. Se observaron 385 y 464 casos de infecciones bacterianas durante los años 2012 y 2013, respectivamente. Acinetobacter baumannii, Escherichia coli y Klebsiella pneumoniae fueron las especies más prevalentes. El consumo total de antibióticos en el Hospital Universitario (HU-FURG), en este período, fue de 3,865 unidades / dosis, siendo 1,995 unidades / dosis en 2012 y 1,870 unidades / dosis en 2013. La relación entre las tasas de los principales patógenos resistentes aislado y se observó el consumo anual del antibiótico correspondiente (carbapenémicos, cefalosporinas, sulfonamidas y otras β-lactamas); Sin embargo, no hubo una asociación significativa entre el aumento de la resistencia y el consumo. Conclusión. Este estudio revela la necesidad de difundir y devolver estos datos a los médicos y tomadores de decisiones en el hospital, ya que puede ser crucial para mejorar las políticas en la prescripción de antibióticos y en la implementación de un control efectivo de infecciones. Introdução. O aumento da produção de novas drogas emerge paralelamente com a adaptação bacteriana a condições adversas. Existem diversos fatores que tem sido apontados como causa desta condição tais como: globalização, uso abusivo de antibióticos, e a prescrição arbitrátia destas drogas. Com isso, o objetivo do estudo foi avaliar o uso de antimicrobianos com a antibiótico-resistência em 2012 e 2013 em um Hospital Universitario do Sul do Brasil. Material e métodos. Este estudo foi realiado no Hospital Universitário Dr. Miguel Riet Correa Jr. em Rio Grande/RS, Brasil. Os dados foram coletados entre Janeiro de 2012 e Dezembro de 2013, na clínica médica e duas unidades de tratamento intensivo. Resultados. Observou-se 385 e 464 casos de infecções bacterianas, durante os anos de 2012 e 2013, respectivamente. Acinetobacter baumannii, Escherichia coli e Klebsiella pneumoniae foram as espécies mais prevalentes. O consumo total de antibióticos no Hospital Universidtário (HU-FURG), neste período, foi de 3.865 unidades/doses, sendo 1.995 unidades/dose em 2012 e 1.870 unidades/dose no ano de 2013. A relação entre as taxas dos principais patógenos resistentes isolados e o consumo anual do antibioticoa correspondentea (carbapenêmicos, cefalosporinas, sulfonamidas e outros β-lactâmicos) foi observada; entretanto, não mostrou associação significativa entre o aumento da resistência e o consumo. Conclusão. Este estudo revela a necessidade de disseminação e retorno destes dados para os clínicos e tomadores de decisão no hospital, pois pode ser crucial o melhoramento de politicas na prescrição de antibiotiocs e na implementação de um crontrole eficaz das infecções..

The combinatorial chemistry has been an important tool for the development of new strategies against the Mycobacterium tuberculosis. Therefore, we evaluated the antimycobacterial activity of two coordinated metal complexes (Cu(II) and Co(II)) and a free ligand, including in the intramacrophage environment. The complexes were more active than the free ligand, indicating that the complexation favoured the antimicrobial activity. None of the compounds showed cytotoxic effect at the concentration of 200 µg ml-1 and both complexes showed intracellular antimicrobial activity, with results as effective as rifampicin. In this study, it was possible to identify complexes containing benzohydroxamate associated with transition metal ions (Cu2+ and Co2+ ), which were able to inhibit the growth of M. tuberculosis, including in persistence stage. In addition, the docking analysis allows inferring a possible interaction of the metal complexes with the enzyme urease, which has been reported as crucial for the bacillus survival in the intraphagosomal environment. Thus, these set of results demonstrate the potential of these metals in the development of new drugs against M. tuberculosis. SIGNIFICANCE AND IMPACT OF THE STUDY: In this study, it was possible to identify complexes containing benzohydroxamate associated with transition metals (Cu2+ and Co2+ ), which were able to inhibit the growth of Mycobacterium tuberculosis, including in the persistence stage. In this context, cobalt and copper can be scaffolds for new drugs against M. tuberculosis.

New drugs are needed to treat infections with antimicrobial-resistant Mycobacterium abscessus; therefore, we evaluated usnic acid as an antimicrobial agent and efflux inhibitor (EI) against M. abscessus. Usnic acid showed antimicrobial activity, and synergistically, the EI verapamil increased this activity. In addition, when we evaluated the interaction of antimicrobials with usnic acid, the increase of their activity was observed. Finally, usnic acid showed an efflux inhibitory effect between the classical EIs verapamil and carbonyl cyanide m-chlorophenylhydrazine. In conclusion, usnic acid showed both antimicrobial and EI activity, indicating that this natural compound may be a promising scaffold for new drugs against this difficult-to-treat microorganism.

Objectives The aims of this study were (i) to determine the frequency of plasmid-mediated resistance to fluoroquinolones (FQs) in Escherichia coli isolated from patients with urinary tract infections (UTIs) of nosocomial and community origin and (ii) to determine the relationships between the presence of extended-spectrum β-lactamases (ESBLs), mutations in the gyrA and parC genes, and resistance to FQs. Methods A total of 71 E. coli isolates, including 38 ESBL-producers and 33 non-ESBL-producers, were analysed. The aac(6′)-Ib gene was amplified using PCR and was subsequently digested with the BtsCI restriction enzyme to identify aac(6′)-Ib-cr, a variant associated with FQ resistance. Detection of qnr genes was performed by multiplex PCR. In isolates that tested positive for these genes, the gyrA and parC genes were sequenced and the modulation factor of an efflux pump inhibitor was determined on the minimum inhibitory concentration (MIC) of norfloxacin. Results The frequencies of qnrS, qnrB and qnrA were 4.2%, 2.8% and 0%, respectively. The frequency of aac(6′)-Ib-cr was 40.8% and this variant was associated with double mutations in gyrA and parC as well as resistance to FQs and ESBL production. Modulation of efflux pump activity was more frequent in resistant isolates that had a wild-type parC gene. Conclusion An interplay of resistance mechanisms increased the level of resistance to FQs, and the high frequency of putative plasmid-mediated quinolone resistance genes associated with ESBL-producing isolates reduced therapeutic options to treat UTIs in the affected population.

Resumen: Introducción: En casos de faringitis por estreptococo grupo A (EGA) se recomienda una pauta de 10 días de amoxicilina. No obstante, parece que pautas de menor duración resultan igualmente efectivas. El objeto de este estudio fue evaluar y comparar de manera retrospectiva la evolución de pacientes tratados con pautas de amoxicilina de 7 y 10 días de duración. Materiales y métodos: Análisis retrospectivo de todos los casos de faringitis por EGA atendidos en un servicio de urgencias en 2014. Se analizaron variables demográficas, uso y resultados de pruebas de detección rápida de antígeno (PDRA), tratamiento, complicaciones y reingreso en un plazo de 30 días. Se definieron 2 grupos para el análisis comparativo basados en la duración del tratamiento con amoxicilina: a) pauta corta (hasta 7 días), y b) pauta larga (10 días). Resultados: Se incluyó a 989 casos de faringitis por EGA. La edad mediana fue 5,2 años, el 50,1% fue de sexo masculino. La amoxicilina fue el antibiótico más prescrito (94,9%), con una duración media de 7 días. Se prescribieron pautas de 10 días al 31,9% de los pacientes. No se encontraron diferencias entre los grupos con pautas cortas y largas en cuanto a la edad (p = 0,600), el género (p = 0,429) o las complicaciones (p = 0,436). Concluimos que en lo referente a la variable de resultado «regreso al servicio de urgencias», la pauta de 7 días no es inferior a la de 10 días. Conclusión: El antibiótico prescrito con mayor frecuencia fue la amoxicilina, aunque solo se prescribió pauta de 10 días en unos pocos casos. Nuestro análisis no encontró beneficio aparente del tratamiento de larga duración con amoxicilina en casos de faringitis por EGA. Abstract: Introduction: In group A streptococcal (GAS) pharyngitis a ten-day course of amoxicillin is recommended. However, short-course treatments seem to be equally effective. The aim of this study was to retrospectively evaluate and compare the outcome of patients treated with 7-day course and 10-day course of amoxicillin. Materials and methods: Retrospective analysis of all GAS pharyngitis admitted to a paediatric emergency department in 2014. Demographic variables, the application and results of the rapid antigenic diagnostic test (RADT), treatment, complications and return in the next 30 days were analysed. Two groups were defined for comparative analysis according to the duration of treatment with amoxicillin: A) short-course (up to 7 days) and B) long-course (10 days). Results: Were included 989 GAS pharyngitis. The median age was 5.2 years, 50.1% male. Amoxicillin was the most prescribed antibiotic (94.9%) with a median duration of 7 days. 10-day course therapy was prescribed in 31.9% of the cases. There were no differences between short and long-course treatment groups regarding age (P=.600), gender (P=.429) and complications (P=.436). Considering the endpoint “return to the emergency department”, we concluded that up to 7 days of treatment was non-inferior to 10 days of treatment. Conclusion: The most commonly prescribed antibiotic was amoxicillin, but a 10-day course was prescribed in few cases. In our analysis there seems to be no benefit with long-course treatments with amoxicillin in GAS pharyngitis.

The objectives of this study were to evaluate tetrahydropyridine derivatives as efflux inhibitors and to understand the mechanism of action of the compounds by in silico studies. Minimum inhibitory concentration (MIC) determination, fluorometric methods and docking simulations were performed. The compounds NUNL02, NUNL09 and NUNL10 inhibited efflux, and NUNL02 is very likely a substrate of the transporter protein AcrB. Docking studies suggested that the mechanism of action could be by competition with substrate for binding sites and protein residues. We showed for the first time the potential of tetrahydropyridines as efflux inhibitors and highlighted compound NUNL02 as an AcrB-specific inhibitor. Docking studies suggested that competition is the putative mechanism of action of these compounds.

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The basis of drug resistance in Mycobacterium abscessus is still poorly understood. Nevertheless, as seen in other microorganisms, the efflux of antimicrobials may also play a role in M. abscessus drug resistance. Here, we investigated the role of efflux pumps in clarithromycin resistance using nine clinical isolates of M. abscessus complex belonging to the T28 erm(41) sequevar responsible for the inducible resistance to clarithromycin. The strains were characterized by drug susceptibility testing in the presence/absence of the efflux inhibitor verapamil and by genetic analysis of drug-resistance-associated genes. Efflux activity was quantified by real-time fluorometry. Efflux pump gene expression was studied by RT-qPCR upon exposure to clarithromycin. Verapamil increased the susceptibility to clarithromycin from 4- to &ge, 64-fold. The efflux pump genes MAB_3142 and MAB_1409 were found consistently overexpressed. The results obtained demonstrate that the T28 erm(41) polymorphism is not the sole cause of the inducible clarithromycin resistance in M. abscessus subsp. abscessus or bolletii with efflux activity providing a strong contribution to clarithromycin resistance. These data highlight the need for further studies on M. abscessus efflux response to antimicrobial stress in order to implement more effective therapeutic regimens and guidance in the development of new drugs against these bacteria.

The treatment for trichomoniasis, based on 5′-nitroimidazol agents, has been presenting failures related to allergic reactions, side effects, and the emergence of resistant isolates. There are no alternative drugs approved for the treatment of these cases; thus, the search for new active molecules is necessary. In this scenario, chalcones have been extensively studied for their promising biological activities. Here, we presented the synthesis of three hydroxychalcones (3a, b, and c), in vitro and in silico analyses against Trichomonas vaginalis. The in vitro biological evaluation showed that hydroxychalcone 3c presented anti-T. vaginalis activity, with complete death in 12 h of incubation at minimum inhibitory concentration (MIC) of 100 μM. 3c showed a dose-dependent cytotoxicity against mammalian VERO cell line, but the association of 3c at 12.5 μM and metronidazole (MTZ) at 40 μM showed 95.31% activity against T. vaginalis trophozoites after 24 h of exposure and did not affect the VERO cell growth, appearing to be a good alternative. In silico analysis by molecular docking showed that 3c could inhibit the activity of TvMGL (methionine gamma-lyase), TvLDH (lactate dehydrogenase), and TvPNP (purine nucleoside phosphorylase) affecting the T. vaginalis survival and also suggesting a different mechanism of action from MTZ. Therefore, these results propose that hydroxychalcones are promising anti-T. vaginalis agents and must be considered for further investigations regarding trichomoniasis treatment.

This study aims to perform a bioactive analysis of five mushrooms collected in south of Brazil. The total phenol content of the extracts was equivalent to the antioxidant activity by ACAP assay. All extracts were able to inhibit the growth of Acinetobacter baumanni, and Auricularia auricula and Lactarius deliciosus extract showed the best antibacterial activity. In addition, no extract showed cytotoxic activity against VERO cells at the highest concentration evaluated (2500 µg/mL). Our results showed better antioxidant activity through the inhibition of the oxidation via peroxyl radical. It can be observed that all extracts were active against A. baumanni, and even moderately, all extracts could be inhibited of at least one of the bacteria used in the study. Added for these, the aqueous extracts showed no toxicity in VERO cells, highlighting the importance of research about the active compounds of mushrooms of the region.

Isoniazid-naphthoquinone hybrids were synthesized and evaluated against a susceptible (H37Rv) strain and two isoniazid-resistant strains (INHR1 and INHR2) of Mycobacterium tuberculosis. The antimycobacterial activity of the derivatives was determined based on the resazurin microtiter assay and their cytotoxicity in adhered mouse monocyte macrophage J774.A1 cells (ATCC TIB-67). Of the twenty-two compounds evaluated against the three strains of M. tuberculosis, twenty-one presented some activity against the H37Rv and INHR1 (katG S315T) or INHR2 (inhA C(−5)T) strains. Compounds 1a, 2a, and 8a were effective against the INHR1 strain, and compounds 1a, 1b, 2a, 3a, 5a, 5b and 8a were effective against the INHR2 strain, with MICs in the range of 3.12–6.25 µg/mL. Compounds 1b and 5b were the most active against H37Rv, with MIC of 0.78 µg/mL. Based on the selectivity index, 1b and 5b can be considered safe as a drug candidate compounds. These results demonstrate that quinoidal compounds can be used as promising scaffolds for the development of new anti-TB drugs and hybrids with activity against M. tuberculosis-susceptible and INH-resistant strains.

Classificada como pandemia pela Organização mundial da saúde, a COVID-19 tornou-se protagonista quanto às demandas de saúde pública ao redor do mundo. No Brasil, segundo país com mais mortes pela doença, o Rio Grande do Sul é o sexto estado mais afetado pelo novo coronavírus. Assim, realizou-se uma pesquisa online com os moradores da cidade de Rio Grande, no sul do estado, para avaliar o impacto da pandemia sobre a população local em relação aos hábitos comportamentais de diferentes grupos. Através de um questionário online foi possível coletar dados de uma amostra de 1188 indivíduos e subsidiar um trabalho quantitativo e descritivo acerca das mudanças comportamentais dos rio-grandinos frente à COVID-19. Os resultados mostraram que apesar de os homens terem sido aqueles que mais mudaram hábitos comportamentais com a chegada da pandemia - como a lavagem frequente das mãos -, as mulheres foram as que tiveram os maiores percentuais de distanciamento social e apresentaram maiores preocupações quanto a serem infectadas. Essa tendência comportamental se reflete nos dados epidemiológicos da cidade, em que mulheres foram minoria em relação ao número de casos e, também, número de óbitos no município. Fatores como renda e escolaridade também influenciam o comportamento dos rio-grandinos em relação aos hábitos durante a pandemia. Dessa forma, o estudo e a compreensão das práticas adotadas pela população é fundamental para que se aperfeiçoem as campanhas preventivas e haja uma readequação no diálogo com a população para aprimorar as medidas de contenção da pandemia.

Justificativa e objetivos: As fluoroquinolonas representam uma classe de antimicrobianos que é frequentemente prescrita no tratamento de infecções do trato urinário (ITU), tanto de origem hospitalar como originárias da comunidade. Este estudo tem como objetivo,determinar a frequência de resistência à ciprofloxacina e levofloxacina, além de seus fatores associados em bacilos Gram negativos (BGN) isolados de pacientes com ITU em um hospital no Sul do Brasil. Métodos: foi realizado um estudo transversal e analítico, baseado em casos de infecções do trato urinário causadas por bactérias gram-negativas provenientes de pacientes atendidos no Hospital Universitário Dr. Miguel Riet Correa Jr. em Rio Grande/RS, de agosto de 2012 a julho 2013. Foram analisadas variáveis independentes como idade, sexo, origem da infecção e características dos isolados clínicos bacterianos. Resultados:Dos 562 BGN isolados e analisados, foi encontrado uma frequência de resistência à ciprofloxacina e à levofloxacina de 25,5 e 23,3%, respectivamente, 62,6% de origem comunitária e 59% de origem hospitalar. Os fatores de risco associados à resistência a ciprofloxacina e levofloxacina, foram pacientes do gênero masculino, infecções adquiridas no ambiente hospitalar, commaior tempo de internação, e apresença de β-lactamases de espectro estendido (ESBL) nos isolados clínicos. Conclusões:Os resultados mostraram uma forte associação da resistência bacteriana de BGN com a permanência no ambiente hospitalar e a presença de ESBL. A fim de controlar a resistência antimicrobiana e a redução nos custos em saúde pública torna-se necessário que os hospitais tenham uma forte política de vigilância contínua do uso e da resistência de antibióticos. Background and Objectives: Fluoroquinolones represent a class of antimicrobial agents that is often prescribed to treat urinary tract infections (UTI), both nosocomial ones and those originating in the community. This study aims to determine the frequency of resistance to ciprofloxacin and levofloxacin and its associated factors in Gram-negative bacilli (GNB) isolated from patients with urinary tract infection in a hospital in southern Brazil. Methods: A cross-sectional analytical study was carried out, based on cases of urinary tract infections caused by gram-negative bacteria in patients treated at Hospital Universitário Dr. Miguel Riet Correa Jr. in Rio Grande, state of Rio Grande do Sul, Brazil, from August 2012 to July 2013. Independent variables such as age, gender, infection origin and characteristics of the clinical and bacterial isolates were analyzed. Results: The resistance rate to ciprofloxacin and levofloxacin was 25.5 and 23.3%, respectively in the 562 BGN isolates that were analyzed, of which 62.6% were originated in community and 59% were nosocomial infections. Risk factors associated with resistance to ciprofloxacin and levofloxacin were male gender, infections acquired in the hospital environment, those with longer hospital stay, and the presence of extended spectrum β-lactamase (ESBL) in the clinical isolates. Conclusions: The results showed a strong association of GNB bacterial resistance with the permanence at the hospital environment and the presence of ESBL. In order to control antimicrobial resistance and reduce public health costs, it becomes necessary for hospitals to have a strong and continuous surveillance policy of antibiotic use and resistance.

Objectives This study aimed to evaluate a tetrahydropyridine derivative (THP) as a potential inhibitor of the efflux mechanism and modulator of the high level of antimicrobial resistance usually observed in members of the Mycobacterium abscessus (M. abscessus) group. Methods The strain M. abscessus subsp. abscessus (ATCC 19997) was used as reference, in addition to three clinical isolates: M. abscessus subsp. abscessus (AT 07), and two M. abscessus subsp. bolletii (AT 46 and AT 52). The minimum inhibitory concentration (MIC) of amikacin (AMI), ciprofloxacin (CIP), clarithromycin (CLA), verapamil (VP), and THP derivative (NUNL02) was determined. Results The NUNL02 showed activity against M. abscessus; the MIC of AMI against ATCC 19997 was reduced more than 16-fold, and the MIC of CIP against AT 52 was reduced four-fold. When combined with CLA, the MIC was reduced against all tested strains. In addition, to detect and quantify the activity of the efflux mechanism, the intracellular accumulation kinetics of the fluorometric substrate ethidium bromide in the presence and absence of VP and NUNL02 were evaluated. The NUNL02 was found to be a more effective efflux inhibitor than VP, which is the classical inhibitor. Conclusions The tetrahydropyridine derivative, NUNL02, is a promising adjuvant in the treatment of infections caused by M. abscessus.

Despite being a curable disease, tuberculosis (TB) remains a public health problem worldwide mainly due to lengthy treatment, as well as its toxic effects, TB/HIV co-infection and the emergence of resistant Mycobacterium tuberculosis strains. These barriers reinforcing the need for development of new antimicrobial agents, that ideally should reduce the time of treatment and be active against susceptible and resistant strains. Quinones are compounds found in natural sources and among them, the naphthoquinones show antifungal, antiparasitic, and antimycobacterial activity. Thus, we evaluated the potential antimycobacterial activity of six 1,4-naphthoquinones derivatives. We determined the minimum inhibitory concentration (MIC) of the compounds against three M. tuberculosis strains: a pan-susceptible H37Rv (ATCC 27294); one mono-resistant to isoniazid (ATCC 35822); and one mono-resistant to rifampicin (ATCC 35838); the cytotoxicity in the J774A.1 (ATCC TIB-67) macrophage lineage; performed in silico analysis about absorption, distribution, metabolism, and excretion (ADME) and docking sites. All evaluated naphthoquinones were active against the three strains with MIC between 206.6 and 12.5 μM, and the compounds with lower MIC values have also showed low cytotoxicity. Moreover, two naphthoquinones derivatives 5 and 6 probably do not exhibit cross resistance with isoniazid and rifampicin, respectively, and regarding ADME analysis, no compound violated the Lipinski's rule-of-five. Considering the set of findings in this study, we conclude that these naphthoquinones could be promising scaffolds to develop new therapeutic strategies to TB.

Tuberculosis (TB) is the cause of more than one million deaths worldwide, and despite being a curable disease, some factors can make therapy difficult, emphasizing the need for the development of new drugs that may potentiate the action of the classic anti-TB antimicrobials. Naphthoimidazoles show a broad spectrum of biological activities, including antimycobacterial activity. The aim of this study was to evaluate the anti-Mycobacterium tuberculosis activity of nine naphthoimidazoles, alone and combined with isoniazid (INH) and rifampicin (RIF). We evaluated the minimum inhibitory concentration (MIC) of the compounds, the fractional inhibitory concentration of the combinations of the naphthoimidazoles with INH or RIF, and the cytotoxicity of these compounds. Eight compounds showed MICs ranging from 1.56 to 25 μg/mL and the presence of substituents on phenyl groups shown to be essential for antimycobacterial activity. Four compounds showed additivity with both INH and RIF and showed SI values higher than 10, indicating safety. Thus, considering the antimycobacterial activity and the absence of antagonism between naphthoimidazoles and the two main drugs for TB treatment, these compounds could be scaffolds for the development of new anti-TB drugs.

In this study, we evaluated the mutations of Helicobacter pylori associated with resistance to clarithromycin and levofloxacin. Furthermore, based on the proposed interaction between antimicrobial resistance and pathogenicity, we correlated the mutation profiles of the strains with the presence of the pathogenicity gene cagA. We analyzed 80 gastric biopsy specimens from H. pylori-infected patients for point mutations in the 23S rRNA gene region and in the gyrA gene, which are related to clarithromycin and levofloxacin resistance, respectively, and investigated the presence of the cagA gene in these strains. We observed that in the assayed biopsies, 8.7% (7/80) had mutations in the 23S rRNA gene region at positions 2143 and 2142, while 22.5% (18/80) had mutations in gyrA at codons 87 and 91. Moreover, absence of the CagA-EPIYA pathogenicity factor was observed in 68% (17/25) of resistant samples. The knowledge of the local profile of antimicrobial resistance and the complex interplay involving resistance and pathogenicity can contribute to an appropriate clinical approach.

Background Mycobacterium abscessus causes a wide range of clinical diseases that are difficult to treat. This microorganism is resistant not only to the classical antituberculosis agents but also to most of the antimicrobials that are currently available, resulting in limited therapeutic options and treatment failure. This scenario stresses the need to search for new drugs with activity against M. abscessus. Objective To evaluate in vitro the antimycobacterial activity and cytotoxicity of rifabutin (RFB 1) and ten derivatives (2-11) against M. abscessus ATCC 19977. Method The minimum inhibitory concentration (MIC) of the molecules was determined by the microdilution broth method according to the guideline described in CLSI. The toxicity evaluation was carried in 96-well microplates, using the cell line J774A.1 (ATCC TIB-67). Result From the eleven molecules tested, RFB 1 and RFB 4 were the compounds showing higher activities against M. abscessus, with MICs of 0.9 and 1.0 µM, respectively. The R1 and R2 moieties seem to have deciding influence over the final activity. Furthermore, N-oxide derivatives 9, 10, and 11 were also active against M. abscessus, with MICs of 7.2 µM, 1.8 µM and 3.8 µM, respectively. An explanatory hypothesis for the better activities of compounds RFB 1, RFB 4, RFB 10 and RFB 11 considers the likely hydrogen bonding between ligands and receptor, balancing the global flexibility and interaction energies. RFB 1 and its most effective derivatives were found to be not toxic. Conclusion Besides RFB 1, its derivatives 4, 10 and 11 show potential for clinical development in the M. abscessus treatment.

Herein, we evaluated tetrahydropyridine (THP) compounds (NUNM) as antimicrobials and inhibitors of the efflux mechanism in M. abscessus. subsp. abscessus. The modulation factor (MF) of efflux inhibitors was calculated from the minimum inhibitory concentrations (MICs) of amikacin (AMI), ciprofloxacin (CIP) and clarithromycin (CLA) in the absence and presence of subinhibitory concentrations of the NUNM compounds and canonical inhibitors carbonyl cyanide m-chlorophenyl hydrazone (CCCP) and verapamil (VP). The kinetics of the intracellular accumulation of the fluorimetric substrate ethidium bromide (EtBr) was evaluated and calculated by the relative final fluorescence (RFF). In addition, molecular modeling simulations for the MmpL5 and Tap efflux transporters with ligands (CLA, NUNM, CCCP, VP and EtBr) were performed to better understand the efflux mechanism. We highlight the NUNM01 compound because it reduced the MICs of AMI, CIP and CLA by 4-, 4- and 16-fold, respectively, had the highest effect on EtBr accumulation (RFF = 3.1) and showed a significant in silico affinity for the evaluated proteins in docking simulations. Based on the analyses performed in vitro and in silico, we propose that NUNM01 is a potential pharmacophore candidate for the development of a therapeutic adjuvant for M. abscessus infections.

Ethnopharmacological relevance Jatropha curcas L. is a plant with high cultural significance for quilombola communities of Oriximina (Para State, Brazil). Although the plant is highly toxic, its seeds are used in these communities to treat tuberculosis and related diseases and symptoms. Aim of the study This study was designed to provide a scientific rationale for the traditional detoxification method and use of J. curcas seeds in quilombola communities of Oriximina. Materials and methods J. curcas seeds were manually separated into testa, tegmen, endosperm, and embryo, and then methanolic extracts of each sample were prepared. The traditional preparation of J. curcas seeds consists of a water extract of endosperms that is known as “milk of pinhao-branco”. The content of phorbol esters (PEs) in the extracts was analyzed by High-Performance Liquid Chromatography with Diode-Array Detection (HPLC-DAD). The cytotoxic activity was evaluated in human monocytic cell line THP-1 by Resazurin Reduction Assay, and antimycobacterial activity was assessed by determining Minimal Inhibitory Concentration (MIC) values against H37Rv and BCG strains using the Resazurin Microtiter Assay (REMA). Results The content analysis revealed that the distribution of PEs within the seeds is not homogeneous. High contents were found in tegmens (4.22 ± 0.25–15.52 ± 0.06 mg/g) and endosperms (1.61 ± 0.07–5.00 ± 0.42 mg/g), while concentrations found in testas and embryos were all below 0.5 mg/g. The traditional preparation derived from the endosperm of J. curcas contained significantly less PEs than the endosperms (0.01 ± 0.005 mg/g). Against THP-1 cells, all the parts of the seed showed cytotoxic activity, while the traditional preparation was considered non-cytotoxic. Nevertheless, only the tegmen and endosperm of J. curcas were considered active against M. tuberculosis and M. bovis (MIC = 200 μg/mL). Conclusion The results of this study indicated that the traditional processing performed by the quilombola people from Oriximina is effective in reducing the toxicity of J. curcas seeds. Although inactive against mycobacteria, the extensive use of the traditional preparation and its low toxicity encourage further studies to investigate other biological activities.

In this paper we report the synthesis and characterization of the following aromatic hydroxamates: potassium(I) benzohydroxamate (BZH), potassium(I) nitrobenzohydroxamate (N-BZH), potassium(I) dinitrobenzohydroxamate (DN-BZH) and the complexes derived containing zinc(II): BZH-Zn, N-BZH-Zn and DN-BZH-Zn. The compounds were characterized and their probable structures proposed using infrared spectroscopy, nuclear magnetic resonance spectroscopy, thermogravimetry and elemental analysis. Calculations of HOMO and LUMO pointed out that DN-BZH anion is the most reactive specie, once it presented the lowest value of energy gap, ca. of 7.65 eV. The compounds were evaluated against Streptococcus pneumoniae, Escherichia coli, Staphylococcus aureus and Pseudomonas aeruginosa. It was possible to observe that DN-BZH demonstrated a minimum inhibitory concentration (MIC) of 25 µg/mL, while its zinc(II) complex presented a MIC of 12.5 µg/mL for S. pneumoniae. Molecular docking studies performed with an urease model suggested coordination between the hydroxamic fraction of DN-BZH and nickel(II) ions located in the active site, showing a bond length of approximately 2.3 A and a variety of interactions with specific amino acids residues.

Background and Objectives: Fluoroquinolones represent a class of amtimicrobials which isfrequently prescribed for the treatment of urinary tract infections (UTIs) of both hospital and community origin. This study aims, to determine the frequency and the factors associated with resistance to ciprofloxacin and levofloxacin in gram-negative bacilli isolated from patients with UTIs treated in a hospital in southern Brazil. Methods: It was performed a transversal and analytical study based on cases of urinary tract infection caused by gram-negative bacteria (GNB) from patients at the Hospital Universitário Dr. Miguel Riet Correa Jr. in Rio Grande/RS from August 2012 to July 2013. Independent variables such as the age and sex of patients, source of infection of the UTI and the characteristics of the bacterial isolates were analyzed. Results: Of the 562 GNB isolated andanalyzed, the total frequencies of ciprofloxacin and levofloxacin resistance were, respectively, 25.5% and 23.3%, 62.6% community origin and 59% in hospital origin. The risk factors associated with ciprofloxacin and levofloxacin resistance were male patients, infections acquired in the hospital, longer hospitalization times, and the presence of extended spectrum β-lactamases (ESBLs) in the clinical isolates. Conclusions:The results showed strong association between GNB resistancewith the time spent in the hospital and the presence of ESBLs. To control antibiotic resistance andthe reduction the costs in public health, it is necessary that thehospitals have a strong policy ofcontinues vigilanceof the use of and of the resistance of antibiotics. KEYWORDS: Bacteria. Fluoroquinolones. Cross Infection. Justificativa e objetivos: As fluoroquinolonas representam uma classe de antimicrobianos que é frequentemente prescrita no tratamento de infecções do trato urinário (ITU), tanto de origem hospitalar como originárias da comunidade. Este estudo tem como objetivo,determinar a frequência de resistência à ciprofloxacina e levofloxacina, além de seus fatores associados em bacilos Gram negativos (BGN) isolados de pacientes com ITU em um hospital no Sul do Brasil. Métodos: foi realizado um estudo transversal e analítico, baseado em casos de infecções do trato urinário causadas por bactérias gram-negativas provenientes de pacientes atendidos no Hospital Universitário Dr. Miguel Riet Correa Jr. em Rio Grande/RS, de agosto de 2012 a julho 2013. Foram analisadas variáveis independentes como idade, sexo, origem da infecção e características dos isolados clínicos bacterianos. Resultados:Dos 562 BGN isolados e analisados, foi encontrado uma frequência de resistência à ciprofloxacina e à levofloxacina de 25,5 e 23,3%, respectivamente, 62,6% de origem comunitária e 59% de origem hospitalar. Os fatores de risco associados à resistência a ciprofloxacina e levofloxacina, foram pacientes do gênero masculino, infecções adquiridas no ambiente hospitalar, commaior tempo de internação, e apresença de β-lactamases de espectro estendido (ESBL) nos isolados clínicos. Conclusões:Os resultados mostraram uma forte associação da resistência bacteriana de BGN com a permanência no ambiente hospitalar e a presença de ESBL. A fim de controlar a resistência antimicrobiana e a redução nos custos em saúde pública torna-se necessário que os hospitais tenham uma forte política de vigilância contínua do uso e da resistência de antibióticos.

INTRODUCTION: The World Health Organization endorses the BACTEC Mycobacterial Growth Indicator Tube (MGIT)(tm) system as a rapid, sensitive, and specific method to diagnostic of tuberculosis. Here, we compared the performance of this system against Ogawa-Kudoh cultures and microscopy. METHODS: A total of 927 samples were obtained between December 2011 and December 2013 from 652 cases of suspected tuberculosis at the School Hospital of the Federal University of Rio Grande in Brazil. RESULTS: The MGIT system confirmed tuberculosis in more cases in less time. CONCLUSIONS: The MGIT system is an effective tool for early diagnosis of tuberculosis, especially in patients with HIV/AIDS.

JUSTIFICATIVA E OBJETIVO: A Ilex paraguariensis St. Hilaire é conhecida popularmente como mate ou erva-mate, consumida popularmente no sul do Brasil, Paraguai e Argentina. Estudos têm evidenciado atividades bioativas da Ilex paraguariensis, como a atividade antioxidante, antinflamatória, diurética, digestiva, cicatricial e estimulante, conferindo um potencial terapêutico à mesma. Tais atividades bioativas da Ilex paraguariensis podem estar relacionadas com os compostos presentes na planta. Como exemplo a presença de polifenóis, flavonoides, xantinas, aminoácidos, vitaminas e minerais. Devido às informações contraditórias sobre os compostos das plantas que contribuem para a atividade antimicrobiana, o presente trabalho teve por objetivo determinar o potencial antimicrobiano dos extratos hexânico, metanólico e aquoso de Ilex paraguariensis, frente as bactérias gram-positivas e gram-negativas. METODOLOGIA: Para avaliação da atividade antimicrobiana foi realizada a técnica de microdiluição em caldo, frente às cepas Staphylococcus aureus (ATCC 12598), Klebsiella pneumoniae (ATCC 700603), Acinetobacter baumannii (ATCC 19606) e Pseudomonas aeruginosa (ATCC 15442). RESULTADOS: A atividade antimicrobiana de Ilex paraguariensis foi demonstrada pelo extrato hexânico e aquoso nos quais apresentaram atividade antimicrobiana frente a todos os microrganismos estudados. Sendo que, a melhor inibição para o extrato aquoso CMI = 25 µg/mL foi frente ao Staphylococcus aureus, enquanto para o extrato hexanico a CMI 100 µg/mL para a Acinetobacter baumannii. CONCLUSÃO: Portanto, a Ilex paraguariensis é uma fonte de pesquisa para novos agentes antimicrobianos naturais. Estes extratos são possíveis candidatos cuja atividade antimicrobiana poderá contribuir para o tratamento de doenças infecciosas causadas tanto por bactérias gram-positivas quanto negativas.

This paper reports the susceptibility profile to rifabutin (RFB) 1 and six recently synthesized RFB analogs 3 – 8 , of either rifampicin (RFP) susceptible Mycobacterium tuberculosis and resistant clinical isolates from two sources: Mexico and Brazil. Taking into account that about 95% of M. tuberculosis strains resistant to RFP present mutations in the rpoB gene, with some of these mutations being determinant also to RFB resistance, the RFB analogs were screened for activity against a set of known RFP susceptible and resistant strains. N ′-Acetyl-RFB 5 and N ′-(undec-10″-enoyl)-RFB 8 showed the best results, in particular with mutations in the codon 516, 522 and 531 of the rpo B gene, and were therefore selected for in vivo assessment of their efficacy. Studies conducted with tuberculous Balb/C mice previously infected with Ser531Leu mutated clinical isolate, evidenced both 5 and 8 as promoters of a significant decrease on tubercle bacilli burden in lungs associated with lower tissue damage, thus confirming them as good leads for drug discovery. The SAR of the acylated compounds 5 and 8 envisaging the identification of pharmacophore features, highlights the importance of profiling more clearly the chemistry within the molecular aspects for elucidation of the mode of action of RFB and analogs, in relation to mutations in Multidrug-Resistant (MDR) strains.

The development of diagnostic tests which can readily differentiate between vaccinated and tuberculosis-infected individuals is crucial for the wider utilization of bacillus Calmette-Guérin (BCG) as vaccine in humans and animals. BCG_0092 is an antigen that elicits specific delayed type hypersensitivity reactions similar in size and morphological aspects to that elicited by purified protein derivative, in both animals and humans infected with the tubercle bacilli. We carried out bioinformatics analyses of the BCG_0092 and designed a diagnostic test by using the predicted MHC class I epitopes. In addition, we performed a knockout of this gene by homologous recombination in the BCG vaccine strain to allow differentiation of vaccinated from infected individuals. For that, the flanking sequences of the target gene (BCG_0092) were cloned into a suicide vector. Spontaneous double crossovers, which result in wild type revertants or knockouts were selected using SacB. BCG_0092 is present only in members of the Mycobacterium tuberculosis complex. Eight predicted MHC class I epitopes with potential for immunological diagnosis were defined, allowing the design of a specific diagnostic test. The strategy used to delete the (BCG_0092) gene from BCG was successful. The knockout genotype was confirmed by PCR and by Southern blot. The mutant BCG strain has the potential of inducing protection against tuberculosis without interfering with the diagnostic test based on the use of selected epitopes from BCG_0092. Keywords: Gene knockout techniques, Clinical diagnosis, BCG vaccine

Tuberculosis (TB), caused by Mycobacterium tuberculosis, is the world's number one killer among infectious diseases. The search for new naturalproducts that can act as drugs against TB has received increased attention duringthe last years. In this work we describe the isolation and identification of the active antimycobacterial principles of the dichloromethane extract from Lippia lacunosa Mart. & Schauer, Verbenaceae. Compounds were evaluated for their in vitro activity against Mycobacterium tuberculosis (susceptible and rifampicin resistantstrain) using a redox bioassay. From the dichloromethane extract of L. lacunosa leaves, seven methoxy-flavones named cirsimaritin (1), eupatilin (2), eupatorin (3), salvigenin (4), 3'-O-methyl-eupatorin (5), 3',7-dimethoxy-5,6,4'- trihydroxyflavone (6), and 7'-O-methylapigenin (7), and one triterpene, named oleanolic acid (8), were isolated. All compounds were found to display antimycobacterial activity against susceptible strain, with MIC ranging from 25 to 200 µg/mL. None of them was active against rifampicin resistant strain. This is the first report in the antimycobacterial activity of 6-substituted flavones, as well as the first report of the occurrence of these substances in L. lacunosa.

The present study was carried out to investigate the presence of polymorphism in the N-acetyltransferase gene of 41 clinical isolates of Mycobacterium tuberculosis, that were resistant to isoniazid (INH) with no mutations in the hot spots of the genes previously described to be involved in INH resistance (katG, inhA and ahpC). We observed single nucleotide polymorphisms (SNPs) in ten of these, including the G619A SNP in five isolates and an additional four so far un-described mutations in another five isolates. Among the latter SNPs, two were synonymous (C276T, n=1 and C375G, n=3), while two more non-synonymous SNPs were composed of C373A (Leu→Met) and T503G (Met→Arg) were observed in respectively one and two isolates. Molecular modeling and structural analysis based in a constructed full length 3D models of wild type TBNAT (TBNAT_H37Rv) and the isoforms (TBNAT_L125M and TBNAT_M168R) were also performed. The refined models show that, just as observed in human NATs, the carboxyl terminus extends deep within the folded enzyme, into close proximity to the buried catalytic triad. Analysis of tbnat that present non-synonymous mutations indicates that both substitutions are plausible to affect enzyme specificity or acetyl-CoA binding capacity. The results contribute to a better understanding of structure-function relationships of NATs. However, further investigation including INH-sensitive strains as a control group is needed to get better understanding of the possible role of these new mutations on tuberculosis control.

Mutations related to streptomycin resistance in the rpsL and rrs genes are well known and can explain about 70% of this phenotypic resistance. Recently, the gidB gene was found to be associated with low-level streptomycin resistance in Mycobacterium tuberculosis . Mutations in gidB have been reported with high frequency, and this gene appears to be very polymorphic, with frameshift and point mutations occurring in streptomycin-susceptible and streptomycin-resistant strains. In this study, mutations in gidB appeared in 27% of streptomycin-resistant strains that contained no mutations in the rpsL or rrs genes, and they were associated with low-level streptomycin resistance. However, the association of certain mutations in gidB with streptomycin resistance needs to be further investigated, as we also found mutations in gidB in streptomycin-susceptible strains. This occurred only when the strain was resistant to rifampin and isoniazid. Two specific mutations appeared very frequently in this and other studies of streptomycin-susceptible and -resistant strains; these mutations were not considered related to streptomycin resistance, but as a polymorphism. We stratified the strains according to the different phylogenetic lineages and showed that the gidB 16 polymorphism (16G allele) was exclusively present in the Latin American-Mediterranean (LAM) genotype, while the gidB 92 polymorphism (92C allele) was associated with the Beijing lineage in another population. In the sample studied, the two characterized single-nucleotide polymorphisms could distinguish LAM and Beijing lineages from the other lineages.

Tuberculosis is still a major health problem worldwide. Although treatment regimens currently available can cure almost all tuberculosis drug susceptible cases, problems such as the length of treatment, the need for multidrug therapy, the emergence of drug resistance, HIV co-infection and persistent Mycobacterium tuberculosis bacilli, stress the need for new anti-tuberculosis drugs. Strategies to search for new anti-tuberculosis drugs involve: screening libraries of small molecules and natural products or the previous identification of targets crucial to the microorganism and the subsequent design of new molecules. Development of new drugs from known compounds having already shown safety and efficacy is an attractive strategy from the economical, pharmaceutical and clinical point of view. Several derivatives of known molecules and new compounds with different targets have been studied with promising preliminary results. Anti-tuberculosis compounds from natural sources have an enormous potential for the development of new drugs, which have shown not only antimicrobial activity per se but also inhibition of the mechanism of resistance (e.g. efflux pumps) or modulation of the immune response (e.g. macrophage stimulation). If these new drugs are going to have an impact in the treatment of the disease they should ideally be active not only against multiplying microorganisms but also against persistent or dormant bacilli. Due to the complexity of the pathology of M. tuberculosis it is unlikely that a single new drug will be enough. This review will discuss strategies for evaluating drug candidates, new targets, new compounds obtained from synthesis and natural sources, and clinical trials that are currently in progress.

A new one-pot strategy for the synthesis of a series of new N-substituted 3-trifluoroacetyl pyrroles is presented. These compounds were obtained by the reaction of 3-trifluoroacetyl-4,5-dihydrofuran with primary amines, which generated 1,1,1-trifluoro-3-(2-hydroxyethyl)-4-alkylaminobut-3-en-2-one intermediates. In most cases these intermediates were not stable enough to be isolated. Thus, in the same reaction vessel they were directly submitted to oxidation with PCC (Corey's reagent) to furnish 1,1,1-trifluoro-3-(2-ethanal)-4-alkylaminobut-3-en-2-ones, which under reflux underwent intramolecular cyclization to give the desired N-substituted 3-trifluoroacetyl pyrroles, in moderate yields. All of these pyrroles were tested against pan-susceptible Mycobacterium tuberculosis H37Rv and clinical isolates INH- and RMP-resistant strain and some of these compounds showed significant in vitro antimicrobial activity.

Clinical utility of rifabutin 1 (RBT), a potent antibiotic used in multidrug regimens for tuberculosis (TB) as well as for infections caused by Mycobacterium avium complex (MAC), has been hampered due to dose-limiting toxicity. RBT analogs 2-11 were synthesized and evaluated against M. avium 1581 and Mycobacterium tuberculosis susceptible and resistant strains in vitro. A selection of candidates were also assayed against non-replicating persistent (NRP) M. tuberculosis. Subsequent in vivo studies with the best preclinical candidate drugs 5 and 8, in a model of progressive pulmonary tuberculosis of Balb/C mice infected either with H(37)Rv drug-sensible strain or with multidrug resistant (MDR) clinical isolates, resistant to all primary antibiotics including rifampicin, were performed. The results disclosed here suggest that 5 and 8 have potential for clinical application.

Thirty-six plant extracts from the brazilian Atlantic Forest were tested for their antimycobacterial activity against Mycobacterium tuberculosis H37Rv and M. kansasii, using the method REMA in seriate concentrations of 100 to 0.20 µg/mL. Among the thirty six extracts tested, five were active against M. tuberculosis, and three of these extracts also showed activity against M. kansasii. Cytotoxicity test with VERO cells was performed with the five extracts active against M. tuberculosis. Only the extract of Peschiera affinis was identified as non-toxic in the concentration of 100µg/mL. Trinta e seis extratos vegetais originários da Mata Atlântica foram testados quanto à sua atividade antimicobacteriana frente ao M. tuberculosis H37Rv e M. kansasii, utilizando o método REMA em concentrações seriadas de 100 a 0,20 µg/mL. Dentre os trinta e seis extratos testados, cinco mostraram atividade frente ao M. tuberculosis, e destes apenas, três mostraram atividade ao M. kansasii, que apresentou susceptibilidade a outros dez. O teste de citotoxicidade com células VERO foi realizado com os cinco extratos ativos frente ao M. tuberculosis em que identificou-se a não toxicidade em apenas um extrato (Peschiera affinis) na concentração de 100 µg/mL.

Tuberculosis remains as a serious infection disease of worldwide distribution, with high morbidity and mortality, mainly in low socio-economic condition countries. The state of emergency of tuberculosis caused by the resistant and multidrug-resistant (MDR) strains, became the main threat to the tuberculosis treatment and control programs. A fast detection method for the resistant strains will allow the implementation of an adequate treatment and contribute for controlling the dissemination of these resistant strains. This study evaluated the performance of the nitrate reductase assay in solid (NRA-LJ) and liquid (NRA-7H9) media, to determine the susceptibility to first line anti-tuberculosis drugs: isoniazid (INH), rifampicin (RMP), ethambutol (EMB) and streptomycin (SMR). Both methods NRA-LJ and NRA-7H9 were evaluated among 18 strains with a known susceptibility profile. The resazurin microtiter assay (REMA) was performed as a reference method. One hundred percent of accordance was observed between NRA-7H9 and REMA for the four tested drugs. When the NRA-LJ method was compared to REMA, the sensitivity and the specificity to INH, RMP, EMB and SMR were 100%, 100 %, 85.7%, 76.9% and 80%, 100%, 75% and 80%, respectively. From the 57 clinical isolates of M. tuberculosis evaluated by NRA-7H9 and REMA, 56 (98.2%) were sensitive to all antibiotics tested (INH, RMP, EMB and SMR) by the NRA-7H9 method, while three of these strains were resistant to INH by REMA. One strain showed resistance to INH and RMP for both methods, and MIC of 1.0 μg/ml to INH for both methods, while MIC of 1.0 and 2.0 μg/ml to RMP for REMA and NRA-7H9, respectively. The three assays showed a high level of agreement for rapid detection of rifampicin and isoniazid resistance. Regarding rapidness, the detection of color change in the NRA method is within instants as compared to the overnight incubation required for the REMA test. NRA might represent an inexpensive and alternative assay for rapid detection of resistance in low-income countries.

Three crude extracts of Aplysina caissara, a marine sponge endemic to Brazil, were tested against a hepatoma cell line and Mycobacterium tuberculosis. The results demonstrate that all extracts are toxic and capable of inhibiting cellular growth. Additionally, the extracts produced morphological aberrations and inhibited cell attachment to culture substrates. These effects were dose/time dependent. Our results also suggest that reactive oxygen species (ROS) production is not involved in the cytotoxic processes levied by the extracts employed in this study and that active metabolites are likely to be present in the polar fractions of the crude extracts. Finally, our results indicate that all three extracts exhibit a moderate anti-tuberculosis capacity, and that the removal of an extract's lipid fraction appears to diminish this activity.

Bovine tuberculosis (BTB) remains an important economic and zoonotic problem in Latin America. Traditionally, the fight against BTB is initiated by the implementation of routine diagnostic tests for certification of free properties. The diagnosis of BTB can be made by direct and indirect methods, in which we can mention clinical, post mortem, histopathological, immunological, bacteriological and molecular methods. The renewal of scientific interest in tuberculosis in recent year has led to develop and improve methods of diagnosis, prevention, control and eradication of BTB. The aim of this review is to present and discuss different diagnosis methods of BTB. Resumo A tuberculose bovina (BTB) continua sendo um importante problema econômico na América Latina, com potenciais consequências zoonóticas. Tradicionalmente, a luta contra a tuberculose bovina tem sido iniciada pela execução de testes de diagnóstico de rotina para a certificação de propriedades livres da doença. O diagnóstico de BTB pode ser feito através de métodos diretos e indiretos, nos quais podemos citar os métodos clínicos, post mortem, histopatológicos, imunológicos, bacteriológicos e moleculares. A renovação do interesse científico em tuberculose nos últimos anos tem levado à necessidade de desenvolver e melhorar os métodos de diagnóstico, prevenção, controle e erradicação da BTB. O objetivo deste artigo é analisar e discutir sobre os diferentes métodos de diagnóstico de BTB.

Fifteen naphthoquinones, sixteen phenazines and fifteen aryl triazoles were synthesized and evaluated against Mycobacterium tuberculosis. Twenty five substances are reported here for the first time and, among all of the compounds evaluated, six presented MIC (minimal inhibitory concentration) values < 6.25 µg mL-1. These substances are promising antimycobacterial prototypes.

Background: The present work aimed to evaluate the antimycobacterial activity and cytotoxicity of Microcystis aeruginosa toxins, the MC-LR variant and purified extract of [D-Leu1 ] microcystin-LR. Methods: The antimicrobial activity of M. aeruginosa extract and microcystin was evaluated by resazurin microtiter assay against Mycobacterium tuberculosis, M. terrae, M. chelonae and M. kansasii. The cytotoxicity assay was performed by trypan blue exclusion against the HTC cell line. Results: Antimicrobial activity was observed in the hexanic extract of M. aeruginosa (RST 9501 strain) against M. tuberculosis, including sensitive and resistant strains with minimal inhibitory concentrations (MIC) between 1.93 μM and 0.06 μM. The high activity of M. aeruginosa hexanic extract could be attributed to the major presence of the toxins MC-LR and [D-Leu1 ] MC-LR that showed activity at MIC between 53 and 0.42 μM against tested mycobacterial strains. Even at the highest concentration tested, no toxicity of M. aeruginosa extracts was identified against HTC cells. Conclusions: These preliminary results suggest that [D-Leu1 ] MC-LR is a promising candidate for the development of a new antimycobacterial agent

Bovine tuberculosis is a major infectious disease of the cattle. In this study, 85 M. bovis isolates from 162 lymph nodes, obtained from a herd of cattle on a farm in southern Brazil, were evaluated using spoligotyping and VNTR. The strains were grouped into five clusters and five orphans, showing a heterogenic genetic profile, what could represent diverse geographic origins of the introduced cows and/or the frequent movement of cattle between different properties.

Mycobacterium bovis is the main causative agent of animal tuberculosis (TB) and it may cause TB in humans. Molecular typing of M. bovis isolates provides precise epidemiological data on issues of inter- or intra-herd transmission and wildlife reservoirs. Techniques used for typing M. bovis have evolved over the last 2 decades, and PCR-based methods such as spoligotyping and mycobacterial interspersed repetitive unit-variable number tandem repeat (MIRU-VNTR) have been extensively used. These techniques can provide epidemiological information about isolates of M. Bovis that may help control bovine TB by indicating possible links between diseased animals, detecting and sampling outbreaks, and even demonstrating cases of laboratory cross-contamination between samples. This review will focus on techniques used for the molecular typing of M. bovis and discuss their general aspects and applications.

Duroia macrophyllapopularly known as “cabeça-de-urubú,” “apuruí,” or “puruí-grande-da-mata” occurs in the Amazon Forest. Its leaves and branches were collected twice and extracted with dichloromethane and methanol. All extracts were subjected to phytochemical investigation and terpenes and flavonoids were found in all dichloromethane and methanol extracts, respectively. Methanol extracts from both branches (1st collection) and leaves (2nd collection) presented hydrolyzed tannins, yet alkaloids were only detected in the dichloromethane and methanol extracts from branches at the 2nd collection. Phenol compounds were found in both dichloromethane extracts’ collections. The action of every extract was assayed againstMycobacterium tuberculosis(RMPr, H37Rv, and INHr strains), showing that the dichloromethane extract from leaves (1st collection) has the major biological activity, with a MIC of 6.25 μg/mL for the INHr strain, 25.0 μg/mL for the RMPr strain, and ≤6.25 μg/mL for the H37Rv strain. The chromatographic fractioning of the dichloromethane extract from leaves (1st collection) yielded the isolation of two triterpenes: oleanolic and ursolic acids, which were identified by NMR analysis and reported for the first time in theDuroiagenus.

summary When bacteria develop drug-resistant mutations, there is often an associated biological cost; however, some strains can exhibit low- or no-cost mutations. In the present study, a quantitative resazurin reduction assay was used to measure the biological cost of Mycobacterium tuberculosis isolates that contained different mutations in the rpsL, rrs, rpoB, and katG genes, and showed different resistance profiles. Biological costs were determined by comparing the growth curves of drug-resistant isolates with drug-susceptible strains. Some strains, such as those with rpoB mutations other than S531L and strains with mutations in all of the studied genes, grew more slowly than did drug-susceptible strains. However, some strains grew more quickly than drug-susceptible strains, such as those that had only the rpsL K43R mutation. Strains with the mutation katG S315T presented heterogeneous biological costs. When analyzed individually, strains with the mutations rpsL43/katG315, rpoB531, and rpoB531/katG315 grew faster than drug-susceptible strains. The results suggest that some strains with the most common mutations correlated to a high resistance toward streptomycin, isoniazid and rifampicin can grow as well as or better than susceptible strains.

Mycobacterium bovis isolates from an outbreak of bovine tuberculosis in a herd of cattle in Rio de Janeiro, Brazil, were analysed by spoligotyping and variable-number tandem repeat PCR analysis of the mycobacterial interspersed repetitive unit and exact tandem repeats. Molecular typing revealed a high genetic diversity of strains in the herd. The genetic diversity could be explained by the introduction of infected animals from different sources.

Tuberculosis (TB), caused by Mycobacterium tuberculosis, is the world’s number one killer among infectious diseases. The search for new natural products that can act as drugs against TB has received increased attention during the last years. In this work we describe the isolation and identification of the active antimycobacterial principles of the dichloromethane extract from Lippia lacunosa Mart. & Schauer, Verbenaceae. Compounds were evaluated for their in vitro activity against Mycobacterium tuberculosis (susceptible and rifampicin resistant strain) using a redox bioassay. From the dichloromethane extract of L. lacunosa leaves, seven methoxy-flavones named cirsimaritin (1), eupatilin (2), eupatorin (3), salvigenin (4), 3′-O-methyl-eupatorin (5), 3′,7-dimethoxy-5,6,4′- trihydroxyflavone (6), and 7′-O-methylapigenin (7), and one triterpene, named oleanolic acid (8), were isolated. All compounds were found to display antimycobacterial activity against susceptible strain, with MIC ranging from 25 to 200 μg/mL. None of them was active against rifampicin resistant strain. This is the first report in the antimycobacterial activity of 6-substituted flavones, as well as the first report of the occurrence of these substances in L. lacunosa.

Tuberculosis remains a serious public health problem, with nine million cases being reported annually. Treatment with antibiotics is the most effective mechanism to control this disease, although the increase in cases with resistant strains, co-infection with HIV, and the long duration of treatment has established the need to develop new drugs. Here we show the activity of usnic acid against susceptible and resistant Mycobacterium tuberculosis strains and against nontuberculous mycobacteria. Further, we did not identify any contribution of efflux in innate resistance to usnic acid.

The essential oil from Anemia tomentosa var. anthriscifolia showed in vitro activity against Mycobacterium tuberculosis (MIC 100 microg/ml) and therefore was characterized by gas chromatography (GC) and by gas chromatography coupled with mass spectrometry (GC-MS). The major constituents of this essential oil were triquinane sesquiterpenes: silphiperfol-6-ene (14.7%), (-)-epi-presilphiperfolan-1-ol (30.6%), presilphiperfol-7-ene (3.9%), cameroonan-7 alpha-ol (4.4%), prenopsan-8-ol (1.9%) and presilphiperfolan-8-ol (8.3%), suggesting the existence of different chemotypes for this species. The essential oil was fractionated by column chromatography and its major constituent and fractions were assayed against Mycobacterium tuberculosis and M. smegmatis. (-)-epi-Presilphiperfolan-1-ol exhibited an MIC of 120 microg/ml against M. tuberculosis H37Rv.

Objetivo: Identificar microrganismos patogênicos na água do porto sul-brasileiro de Rio Grande, incluindo bactérias, protozoários e fungos. Metodologia: Foram obtidas amostras entre Junho e Novembro de 2006, em pontos representativos ao longo do Porto e analisadas para a presença de bactérias, protozoários e fungos. Foi realizada uma análise qualitativa e quantitativa dos microrganismos. Resultados: Todos os pontos tiveram contagens elevadas de coliformes totais e fecais. Protozoários foram encontrados em 61,9% das amostras representados por Acanthamoeba spp. (28,6%), Giardia spp (16,7%), Criptosporidum spp. (9,5%) e Naegleria spp. (7,1%). Os fungos foram positivos em 88,3% dos casos. Os mais frequentes foram Aspergillus spp. (33,1 %), Penicillium spp.(23,8%) Cladosporium spp. (21,4%) e Rodothorula spp. (11,9%). Conclusões: A água portuária contém não só bactérias patogênicas, mas também protozoários e fungos. Esta situação exige uma vigilância apropriada de forma a reduzir o risco em virtude das atividades de lastreamento e deslastreamento de navios. The study aimed to identify microorganisms in the water of a Brazilian Port, including protozoa and fungus, due to the risk of increasing ballast water activities. Water samples were obtained between June and November 2006, from representative sites of the Port, and analyzed for the presence of bacteria, protozoa and fungus. Qualitative and quantitative counts of microorganisms were performed. Higher counts of total and fecal coliforms were identified in all stations. Protozoa were found in 61.9% of the samples: Acanthamoeba spp. (28.6%), Giardia spp (16.7%), Criptosporidum spp. (9.5%) and Naegleria spp. (7.1%). Fungus were positive in 88.3% of the samples and the most frequent were Aspergillus spp. (33.1%), Penicillium spp.(23.8%) Cladosporium spp. (21.4%), Rodothorula spp. (11.9%). Samples from Brazilian port water contains not only pathogenic bacteria but also protozoa and fungus, which demands appropriate surveillance and measures to reduce the potential hazard in view of deballasting and ballasting ship activities.

A new one-pot strategy for the synthesis of a series of new N-substituted 3-trifluoroacetyl pyrroles is presented. These compounds were obtained by the reaction of 3-trifluoroacetyl-4,5-dihydrofuran with primary amines, which generated 1,1,1-trifluoro-3-(2-hydroxyethyl)-4-alkylaminobut-3-en-2-one intermediates. In most cases these intermediates were not stable enough to be isolated. Thus, in the same reaction vessel they were directly submitted to oxidation with PCC (Corey's reagent) to furnish 1,1,1-trifluoro-3-(2-ethanal)-4-alkylaminobut-3-en-2-ones, which under reflux underwent intramolecular cyclization to give the desired N-substituted 3-trifluoroacetyl pyrroles, in moderate yields. All of these pyrroles were tested against pan-susceptible Mycobacterium tuberculosis H37Rv and clinical isolates INH- and RMP-resistant strain and some of these compounds showed significant in vitro antimicrobial activity.

As doenças infecciosas ainda são uma das principais causas de morte no mundo, sendo de significativa importância o desenvolvimento de novos compostos antimicrobianos contra diferentes microrganismos. As plantas podem ser uma boa fonte para direcionar a busca destes compostos. Neste estudo, 66 extratos de 25 plantas da floresta ribeirinha do sul do Rio Uruguai foram estudados para a atividade antimicrobiana contra o Staphylococcus aureus, Listeria inocua, Escherichia coli, Pseudomonas aeruginosa, Mycobacterium tuberculosis, Aspergillus niger e Candida albicans. Cinqüenta e três destes extratos apresentaram algum tipo de atividade antimicrobiana. Seis (Eugenia mansoni, Eugenia repanda, Myrcianthes cisplatensis, Paullinia ellegans, Petunia sp e Ruprechtia laxiflora) apresentaram atividade contra o Mycobacterium tuberculosis com CIM de 50 μg/mL. Development of new antimicrobial compounds against different microorganisms is becoming critically important, as infectious diseases are still one of the leading causes of death in the world. Plants can be a useful source of these lead compounds. In this study, 66 extracts of 25 plants of the riverside forest of southern Uruguay River were studied for antimicrobial activity against Staphylococcus aureus, Listeria inocua, Escherichia coli, Pseudomonas aeruginosa, Mycobacterium tuberculosis, Aspergillus niger and Candida albicans. Fifty-three of these extracts showed some kind of antimicrobial activity. Six of these (Eugenia mansoni, Eugenia repanda, Myrcianthes cisplatensis, Paullinia ellegans, Petunia sp and Ruprechtia laxiflora)presented activity against Mycobacterium tuberculosis with MIC values as low as 50 μg/mL.