A nucleoid-associated protein is involved in the emergence of antibiotic resistance by promoting the frequent exchange of the replicative DNA polymerase in Mycobacterium smegmatis.

Antibiotic resistance in Mycobacterium tuberculosis exclusively originates from chromosomal mutations, either during normal DNA replication or under stress, when the expression of error-prone DNA polymerases increases to repair damaged DNA. To bypass DNA lesions and catalyze error-prone DNA synthesis, translesion polymerases must be able to access the DNA, temporarily replacing the high-fidelity replicative polymerase. The mechanisms that govern polymerase exchange are not well understood, especially in mycobacteria. Here, using a suite of quantitative fluorescence imaging techniques, we discover that in Mycobacterium smegmatis, as in other bacterial species, the replicative polymerase, DnaE1, exchanges at a timescale much faster than that of DNA replication. Interestingly, this fast exchange rate depends on an actinobacteria-specific nucleoid-associated protein (NAP), Lsr2. In cells missing lsr2, DnaE1 exchanges less frequently, and the chromosome is replicated more faithfully. Additionally, in conditions that damage DNA, cells lacking lsr2 load the complex needed to bypass DNA lesions less effectively and, consistently, replicate with higher fidelity but exhibit growth defects. Together, our results show that Lsr2 promotes dynamic flexibility of the mycobacterial replisome, which is critical for robust cell growth and lesion repair in conditions that damage DNA. IMPORTANCE: Unlike many other pathogens, Mycobacterium tuberculosis has limited ability for horizontal gene transfer, a major mechanism for developing antibiotic resistance. Thus, the mechanisms that facilitate chromosomal mutagenesis are of particular importance in mycobacteria. Here, we show that Lsr2, a nucleoid-associated protein, has a novel role in DNA replication and mutagenesis in the model mycobacterium Mycobacterium smegmatis. We find that Lsr2 promotes the fast exchange rate of the replicative DNA polymerase, DnaE1, at the replication fork and is important for the effective loading of the DnaE2-ImuA'-ImuB translesion complex. Without lsr2, M. smegmatis replicates its chromosome more faithfully and acquires resistance to rifampin at a lower rate, but at the cost of impaired survival to DNA damaging agents. Together, our work establishes Lsr2 as a potential factor in the emergence of mycobacterial antibiotic resistance.

A nucleoid-associated protein is involved in the emergence of antibiotic resistance by promoting the frequent exchange of the replicative DNA polymerase in M. smegmatis.

Antibiotic resistance in M. tuberculosis exclusively originates from chromosomal mutations, either during normal DNA replication or under stress, when the expression of error-prone DNA polymerases increases to repair damaged DNA. To bypass DNA lesions and catalyze error-prone DNA synthesis, translesion polymerases must be able to access the DNA, temporarily replacing the high-fidelity replicative polymerase. The mechanisms that govern polymerase exchange are not well understood, especially in mycobacteria. Here, using a suite of quantitative fluorescence imaging techniques, we discover that, as in other bacterial species, in M. smegmatis, the replicative polymerase, DnaE1, exchanges at a timescale much faster than that of DNA replication. Interestingly, this fast exchange rate depends on an actinobacteria-specific nucleoid-associated protein (NAP), Lsr2. In cells missing lsr2, DnaE1 exchanges less frequently, and the chromosome is replicated more faithfully. Additionally, in conditions that damage DNA, cells lacking lsr2 load the complex needed to bypass DNA lesions less effectively and, consistently, replicate with higher fidelity but exhibit growth defects. Together, our results show that Lsr2 promotes dynamic flexibility of the mycobacterial replisome, which is critical for robust cell growth and lesion repair in conditions that damage DNA.

Spatial segregation and aging of metabolic processes underlie phenotypic heterogeneity in mycobacteria.

Individual cells within clonal populations of mycobacteria vary in size, growth rate, and antibiotic susceptibility. Heterogeneity is, in part, determined by LamA, a protein found exclusively in mycobacteria. LamA localizes to sites of new cell wall synthesis where it recruits proteins important for polar growth and establishing asymmetry. Here, we report that in addition to this function, LamA interacts with complexes involved in oxidative phosphorylation (OXPHOS) at a subcellular location distinct from cell wall synthesis. Importantly, heterogeneity depends on a unique extension of the mycobacterial ATP synthase, and LamA mediates the coupling between ATP production and cell growth in single cells. Strikingly, as single cells age, concentrations of proteins important for oxidative phosphorylation become less abundant, and older cells rely less on oxidative phosphorylation for growth. Together, our data reveal that central metabolism is spatially organized within a single mycobacterium and varies within a genetically identical population of mycobacteria. Designing therapeutic regimens to account for this heterogeneity may help to treat mycobacterial infections faster and more completely.

An essential periplasmic protein coordinates lipid trafficking and is required for asymmetric polar growth in mycobacteria.

Mycobacteria, including the human pathogen Mycobacterium tuberculosis, grow by inserting new cell wall material at their poles. This process and that of division are asymmetric, producing a phenotypically heterogeneous population of cells that respond non-uniformly to stress (Aldridge et al., 2012; Rego et al., 2017). Surprisingly, deletion of a single gene - lamA - leads to more symmetry, and to a population of cells that is more uniformly killed by antibiotics (Rego et al., 2017). How does LamA create asymmetry? Here, using a combination of quantitative time-lapse imaging, bacterial genetics, and lipid profiling, we find that LamA recruits essential proteins involved in cell wall synthesis to one side of the cell - the old pole. One of these proteins, MSMEG_0317, here renamed PgfA, was of unknown function. We show that PgfA is a periplasmic protein that interacts with MmpL3, an essential transporter that flips mycolic acids in the form of trehalose monomycolate (TMM), across the plasma membrane. PgfA interacts with a TMM analog suggesting a direct role in TMM transport. Yet our data point to a broader function as well, as cells with altered PgfA levels have differences in the abundance of other lipids and are differentially reliant on those lipids for survival. Overexpression of PgfA, but not MmpL3, restores growth at the old poles in cells missing lamA. Together, our results suggest that PgfA is a key determinant of polar growth and cell envelope composition in mycobacteria, and that the LamA-mediated recruitment of this protein to one side of the cell is a required step in the establishment of cellular asymmetry.

Mycobacterial serine/threonine phosphatase PstP is phosphoregulated and localized to mediate control of cell wall metabolism.

The mycobacterial cell wall is profoundly regulated in response to environmental stresses, and this regulation contributes to antibiotic tolerance. The reversible phosphorylation of different cell wall regulatory proteins is a major mechanism of cell wall regulation. Eleven serine/threonine protein kinases phosphorylate many critical cell wall-related proteins in mycobacteria. PstP is the sole serine/ threonine phosphatase, but few proteins have been verified as PstP substrates. PstP is itself phosphorylated, but the role of its phosphorylation in regulating its activity has been unclear. In this study, we aim to discover novel substrates of PstP in Mycobacterium tuberculosis (Mtb). We show in vitro that PstP dephosphorylates two regulators of peptidoglycan in Mtb, FhaA, and Wag31. We also show that a phosphomimetic mutation of T137 on PstP negatively regulates its catalytic activity against the cell wall regulators FhaA, Wag31, CwlM, PknB, and PknA, and that the corresponding mutation in Mycobacterium smegmatis causes misregulation of peptidoglycan in vivo. We show that PstP is localized to the septum, which likely restricts its access to certain substrates. These findings on the regulation of PstP provide insight into the control of cell wall metabolism in mycobacteria.

Cell Wall Damage Reveals Spatial Flexibility in Peptidoglycan Synthesis and a Nonredundant Role for RodA in Mycobacteria.

Cell wall peptidoglycan is a heteropolymeric mesh that protects the bacterium from internal turgor and external insults. In many rod-shaped bacteria, peptidoglycan synthesis for normal growth is achieved by two distinct pathways: the Rod complex, comprised of MreB, RodA, and a cognate class B penicillin-binding protein (PBP), and the class A PBPs (aPBPs). In contrast to laterally growing bacteria, pole-growing mycobacteria do not encode an MreB homolog and do not require SEDS protein RodA for in vitro growth. However, RodA contributes to the survival of Mycobacterium tuberculosis in some infection models, suggesting that the protein could have a stress-dependent role in maintaining cell wall integrity. Under basal conditions, we find here that the subcellular distribution of RodA largely overlaps that of the aPBP PonA1 and that both RodA and the aPBPs promote polar peptidoglycan assembly. Upon cell wall damage, RodA fortifies Mycobacterium smegmatis against lysis and, unlike aPBPs, contributes to a shift in peptidoglycan assembly from the poles to the sidewall. Neither RodA nor PonA1 relocalize; instead, the redistribution of nascent cell wall parallels that of peptidoglycan precursor synthase MurG. Our results support a model in which mycobacteria balance polar growth and cell-wide repair via spatial flexibility in precursor synthesis and extracellular insertion. IMPORTANCE Peptidoglycan synthesis is a highly successful target for antibiotics. The pathway has been extensively studied in model organisms under laboratory-optimized conditions. In natural environments, bacteria are frequently under attack. Moreover, the vast majority of bacterial species are unlikely to fit a single paradigm of cell wall assembly because of differences in growth mode and/or envelope structure. Studying cell wall synthesis under nonoptimal conditions and in nonstandard species may improve our understanding of pathway function and suggest new inhibition strategies. Mycobacterium smegmatis, a relative of several notorious human and animal pathogens, has an unusual polar growth mode and multilayered envelope. In this work, we challenged M. smegmatis with cell wall-damaging enzymes to characterize the roles of cell wall-building enzymes when the bacterium is under attack.

Fighting microbial pathogens by integrating host ecosystem interactions and evolution.

Successful therapies to combat microbial diseases and cancers require incorporating ecological and evolutionary principles. Drawing upon the fields of ecology and evolutionary biology, we present a systems-based approach in which host and disease-causing factors are considered as part of a complex network of interactions, analogous to studies of "classical" ecosystems. Centering this approach around empirical examples of disease treatment, we present evidence that successful therapies invariably engage multiple interactions with other components of the host ecosystem. Many of these factors interact nonlinearly to yield synergistic benefits and curative outcomes. We argue that these synergies and nonlinear feedbacks must be leveraged to improve the study of pathogenesis in situ and to develop more effective therapies. An eco-evolutionary systems perspective has surprising and important consequences, and we use it to articulate areas of high research priority for improving treatment strategies.

Loss of functional capacity in elderly individuals with Alzheimer disease.

BACKGROUND: The functional capacity of elderly individuals with Alzheimer disease (AD) progressively declines. OBJECTIVE: To verify the influence of sociodemographic, clinical, staging, mobility, and postural and cognitive balance data on the impairment of the functional capacity of elderly individuals with AD. METHODS: This observational, analytical, cross-sectional study was performed at the Physiotherapy Department of the Universidade Federal do Rio Grande do Norte, Natal, Rio Grande do Norte, Brazil. The study consisted of forty elderly individuals aged ≥60 years old with mild or moderate AD, who could ambulate independently. The instruments used included a questionnaire to assess sociodemographic and anthropometric data; the Mini-Mental Health State Examination (MMSE); the Clinical Dementia Rating (CDR); a clock drawing test (CDT); a verbal fluency test (VFT); the Timed Up and Go Test (TUG); and the Clinical Test of Sensory Organization and Balance (CTSIB). Simple descriptive analyses, Mann-Whitney test, Spearman's correlation test, linear regression modeling, and prediction equation (p<0.05, 95% confidence interval [95%CI]) were performed. RESULTS: Fifteen linear regression models were generated, with the final model chosen for analysis. The variables assumed in that model were CDR, MMSE score, and condition 3 of the CTSIB, which explained 60.1% of the outcome. CONCLUSIONS: Impairment of functional capacity in elderly individuals with AD was influenced by disease progression, which was due to cognitive deficits and deficits in postural balance, which are related to the inaccuracy of the somatosensory system in performing sensory integration.

INTRODUÇÃO: A capacidade funcional de idosos com doença de Alzheimer (DA) sofrerá prejuízo progressivo. OBJETIVO: O presente estudo visou verificar a influências de dados sociodemográficos, clínicos, de estadiamento, mobilidade, equilíbrio postural e cognitivos no prejuízo da capacidade funcional de idosos com DA. MÉTODOS: Trata-se de um estudo observacional, analítico e transversal, realizado no Departamento de Fisioterapia da Universidade Federal do Rio Grande do Norte, em Natal, Rio Grande do Norte, Brasil. O estudo contou com a participação de 40 idosos com idade igual ou superior a 60 anos com DA leve ou moderada, com deambulação independente. Os instrumentos utilizados incluíram um questionário para avaliação de dados sociodemográficos e antropométricos; o Mini-Exame de Estado de Saúde Mental (MEEM); a Avaliação Clínica da Demência (CDR); o Teste do desenho do Relógio (TDR); o Teste de Fluência Verbal (TFV); o Timed Up and Go Test (TUG); e o Clinical Test of Sensory Organization and Balance (CTSIB). Foram realizadas análises descritivas simples, teste de Mann-Whitney, teste de correlação de Spearman, modelo de regressão linear e equação de predição (p<0,05 e intervalo de 95% [IC95%]). RESULTADOS: Foram gerados quinze modelos de regressão linear e foi escolhido o último para a análise. As variáveis assumidas nesse modelo citado foram: CDR, MEEM e condição três do CTSIB, que explicam 60,1% do desfecho. CONCLUSÕES: Pode-se concluir que o prejuízo da capacidade funcional de idosos com DA é influenciado pelo avançar da doença, pelos déficits cognitivo e de equilíbrio postural, sendo esse mais relacionado à imprecisão do sistema somatossensorial em realizar a integração sensorial.

The conserved translation factor LepA is required for optimal synthesis of a porin family in Mycobacterium smegmatis.

The recalcitrance of mycobacteria to antibiotic therapy is in part due to its ability to build proteins into a multi-layer cell wall. Proper synthesis of both cell wall constituents and associated proteins is crucial to maintaining cell integrity, and intimately tied to antibiotic susceptibility. How mycobacteria properly synthesize the membrane-associated proteome, however, remains poorly understood. Recently, we found that loss of lepA in Mycobacterium smegmatis (Msm) altered tolerance to rifampin, a drug that targets a non-ribosomal cellular process. LepA is a ribosome-associated GTPase found in bacteria, mitochondria, and chloroplasts, yet its physiological contribution to cellular processes is not clear. To uncover the determinants of LepA-mediated drug tolerance, we characterized the whole-cell proteomes and transcriptomes of a lepA deletion mutant relative to strains with lepA We find that LepA is important for the steady-state abundance of a number of membrane-associated proteins, including an outer membrane porin, MspA, which is integral to nutrient uptake and drug susceptibility. Loss of LepA leads to a decreased amount of porin in the membrane which leads to the drug tolerance phenotype of the lepA mutant. In mycobacteria, the translation factor LepA modulates mycobacterial membrane homeostasis, which in turn affects antibiotic tolerance.ImportanceThe mycobacterial cell wall is a promising target for new antibiotics due to the abundance of important membrane-associated proteins. Defining mechanisms of synthesis of the membrane proteome will be critical to uncovering and validating drug targets. We found that LepA, a universally conserved translation factor, controls the synthesis of a number of major membrane proteins in M. smegmatis LepA primarily controls synthesis of the major porin MspA. Loss of LepA results in decreased permeability through the loss of this porin, including permeability to antibiotics like rifampin and vancomycin. In mycobacteria, regulation from the ribosome is critical for the maintenance of membrane homeostasis and, importantly, antibiotic susceptibility.

Loss of functional capacity in elderly individuals with Alzheimer disease / Prejuízo da capacidade funcional de idosos com doença de alzheimer

ABSTRACT. Background: The functional capacity of elderly individuals with Alzheimer disease (AD) progressively declines. Objective: To verify the influence of sociodemographic, clinical, staging, mobility, and postural and cognitive balance data on the impairment of the functional capacity of elderly individuals with AD. Methods: This observational, analytical, cross-sectional study was performed at the Physiotherapy Department of the Universidade Federal do Rio Grande do Norte, Natal, Rio Grande do Norte, Brazil. The study consisted of forty elderly individuals aged ≥60 years old with mild or moderate AD, who could ambulate independently. The instruments used included a questionnaire to assess sociodemographic and anthropometric data; the Mini-Mental Health State Examination (MMSE); the Clinical Dementia Rating (CDR); a clock drawing test (CDT); a verbal fluency test (VFT); the Timed Up and Go Test (TUG); and the Clinical Test of Sensory Organization and Balance (CTSIB). Simple descriptive analyses, Mann-Whitney test, Spearman's correlation test, linear regression modeling, and prediction equation (p<0.05, 95% confidence interval [95%CI]) were performed. Results: Fifteen linear regression models were generated, with the final model chosen for analysis. The variables assumed in that model were CDR, MMSE score, and condition 3 of the CTSIB, which explained 60.1% of the outcome. Conclusions: Impairment of functional capacity in elderly individuals with AD was influenced by disease progression, which was due to cognitive deficits and deficits in postural balance, which are related to the inaccuracy of the somatosensory system in performing sensory integration.

RESUMO. Introdução: A capacidade funcional de idosos com doença de Alzheimer (DA) sofrerá prejuízo progressivo. Objetivo: O presente estudo visou verificar a influências de dados sociodemográficos, clínicos, de estadiamento, mobilidade, equilíbrio postural e cognitivos no prejuízo da capacidade funcional de idosos com DA. Métodos: Trata-se de um estudo observacional, analítico e transversal, realizado no Departamento de Fisioterapia da Universidade Federal do Rio Grande do Norte, em Natal, Rio Grande do Norte, Brasil. O estudo contou com a participação de 40 idosos com idade igual ou superior a 60 anos com DA leve ou moderada, com deambulação independente. Os instrumentos utilizados incluíram um questionário para avaliação de dados sociodemográficos e antropométricos; o Mini-Exame de Estado de Saúde Mental (MEEM); a Avaliação Clínica da Demência (CDR); o Teste do desenho do Relógio (TDR); o Teste de Fluência Verbal (TFV); o Timed Up and Go Test (TUG); e o Clinical Test of Sensory Organization and Balance (CTSIB). Foram realizadas análises descritivas simples, teste de Mann-Whitney, teste de correlação de Spearman, modelo de regressão linear e equação de predição (p<0,05 e intervalo de 95% [IC95%]). Resultados: Foram gerados quinze modelos de regressão linear e foi escolhido o último para a análise. As variáveis assumidas nesse modelo citado foram: CDR, MEEM e condição três do CTSIB, que explicam 60,1% do desfecho. Conclusões: Pode-se concluir que o prejuízo da capacidade funcional de idosos com DA é influenciado pelo avançar da doença, pelos déficits cognitivo e de equilíbrio postural, sendo esse mais relacionado à imprecisão do sistema somatossensorial em realizar a integração sensorial.

Perfil nutricional de pacientes onco-hematológico internados em um Hospital Especializado em câncer em São Luís - MA / Nutritional Profile of Onco-hematological Patients Hospitalized in a Cancer Specialized Hospital in São Luís - MA

Objetivo: Avaliar o perfil nutricional de pacientes onco-hematológicos internados em um hospital especializado em câncer em São Luís - MA. Metodologia: Estudo transversal, retrospectivo, analítico, com coleta de dados secundária, envolvendo pacientes com idade mínima de 18 anos, de ambos os gêneros e que tenham sido submetidos a pelo menos uma ASG-PPP (Avaliação Subjetiva Global Produzida Pelo Paciente). Os dados foram coletados em registros do Serviço de Nutrição e Dietética do hospital. Analisaram-se dados demográficos (gênero e idade), clínicos (diagnóstico) e nutricionais: Índice de massa corporal (IMC), circunferên-cia braquial (CB), prega cutânea tricipital (PCT), circunferência muscular do braço (CMB) e ASG-PPP. As análises foram realizadas no programa estatístico Stata® 13.0. O nível de significância utilizado para os testes foi de p<0,05. Resultados: Foram avaliados 330 pacientes, onde foi constatado que 67,58% eram de adultos e 32,42% de idosos, com maior frequência do sexo masculino, com 60,30%. Ocorreu maior incidência de leucemia (58,48%), seguidos de linfomas (24,85%), mieloma múltiplo (13,33%) e síndrome mielodisplásica (3,3%). Na avaliação do estado nutricional os resultados mostraram que a ASG-PPP detectou maior número de pacientes com algum grau de desnutrição do que outros indicadores (93,94%), seguido pela PCT (65,76%), CMB (53,64%), CB (45,45%) e IMC (14,87%). De acordo com o IMC, foi encontrado maior incidência de eutrofia, correspondendo a 57,27% da amostra. Conclu-são: Diante do que foi encontrado, destaca-se que a desnutrição é um aspecto de extrema importância a ser considerado no tratamento de pacientes onco-hematológicos, visto que pode interferir diretamente no prognóstico da doença. (AU)

Objective: To evaluate the nutritional profile of onco-hematological patients admitted to a specialized cancer hospital in São Luís - MA. Methodology: Cross-sectional, retrospective, analytical study, with secondary data collection, involving patients aged at least 18 years, of both genders and having undergone at least one ASG-PPP (Subjective Global Assessment Produced by the Patient). Data were collected from records of the Hospital's Nutrition and Dietetics Service. Demographic (gender and age), clinical (diagnostic) and nutritional: Body Mass Index (BMI), Brachial Circumference (CB), Tricipital Skinfold (PCT), Muscular Arm Circumference (CMB) and ASG-PPP data were analyzed. The analyses were performed using the Stata® 13.0 statistical program. The level of significance used for the tests was p<0.05. Results: 330 patients were evaluated, in which it was found that 67.58% are adults and 32.42% are elderly, with a higher frequency of males with 60.30%. There was a higher incidence of Leukemia (58.48%), followed by Lymphomas (24.85%), Multiple Myeloma (13.33%) and Myelodysplastic Syndrome (3.3%). In the assessment of nutritional status, the results showed that ASG-PPP detected a greater number of patients with some degree of malnutrition than other indicators (93.94%), followed by PCT (65.76%), CMB (53.64%), CB (45.45%) and BMI (14.87%). According to the BMI, a higher incidence of eutrophy was found, which corresponds to 57.27% of the sample. Conclusion: In view of what was found, it is highlighted that malnutrition is an extremely important aspect to be considered in the treatment of onco-hematological patients, since it can directly interfere in the prognosis of the disease. (AU)

Itaconate is an effector of a Rab GTPase cell-autonomous host defense pathway against Salmonella.

The guanosine triphosphatase (GTPase) Rab32 coordinates a cell-intrinsic host defense mechanism that restricts the replication of intravacuolar pathogens such as Salmonella Here, we show that this mechanism requires aconitate decarboxylase 1 (IRG1), which synthesizes itaconate, a metabolite with antimicrobial activity. We find that Rab32 interacts with IRG1 on Salmonella infection and facilitates the delivery of itaconate to the Salmonella-containing vacuole. Mice defective in IRG1 rescued the virulence defect of a S. enterica serovar Typhimurium mutant specifically defective in its ability to counter the Rab32 defense mechanism. These studies provide a link between a metabolite produced in the mitochondria after stimulation of innate immune receptors and a cell-autonomous defense mechanism that restricts the replication of an intracellular bacterial pathogen.

LIVER TRANSPLANTATION FOR BILE DUCT INJURY AFTER CHOLECYSTECTOMY.

BACKGROUND: Bile duct injury is a life-threatening complication that requires proper management to prevent the onset of negative outcomes. Patients may experience repeated episodes of cholangitis, secondary biliary cirrhosis, end-stage liver disease and death. OBJECTIVE: To report a single center experience in iatrogenic secondary liver transplantation after cholecystectomy and review the literature. METHODS: This was a retrospective single center study. Of the 1662 liver transplantation realized, 10 (0.60 %) were secondary to iatrogenic bile ducts injuries due cholecystectomies. Medical records of these patients were reviewed in this study. RESULTS: Nine of 10 patients were women; the median time in waiting list and between cholecystectomy and inclusion in waiting list was of 222 days and of 139.9 months, respectively. Cholecystectomy was performed by open approach in eight (80%) cases and by laparoscopic approach in two (20%) cases. The patients underwent an average of 3.5 surgeries and procedures before liver transplantation. Biliary reconstruction was realized with a Roux-en-Y hepaticojejunostomy in nine (90%) cases. Mean operative time was 447.2 minutes and the median red blood cell transfusion was 3.4 units per patient. Mortality in the first month was of 30%. CONCLUSION: Although the liver transplantation is an extreme treatment for an initially benign disease, it has its well-defined indications in treatment of bile duct injuries after cholecystectomy, either in acute or chronic scenario.

Toxoplasmosis outbreak in Brazil, 2006 - Revisited.

Waterborne outbreaks of human toxoplasmosis can have great magnitude due to the number of persons infected while smaller-scale outbreaks are also possible. This is a study based on a historical database investigating a toxoplasmosis outbreak occurred in 2006 in a residential community in São Luís, in the Brazilian state of Maranhão. Ninety of the 110 residents, employees and domestic helping persons had blood samples collected and tested. The diagnosis of toxoplasmosis was established by quantification of anti-Toxoplasma gondii immunoglobulin M and immunoglobulin G antibodies using enzyme immunoassay. The subjects were classified as past infection, acute/recent infection or seronegatives. The definition of acute infection was based on the presence of indicative symptoms and immunoglobulin M positivity. There were 33 cases of acute infection. The outbreak was concluded to be waterborne: consumption of faucet-mount filtered water was indicated as risk factor. We discuss the challenges of investigating waterborne toxoplasmosis outbreaks.

Transplante de fígado para tratamento de lesão de via biliar após colecistectomia / Liver transplantation for bile duct injury after cholecystectomy

ABSTRACT BACKGROUND: Bile duct injury is a life-threatening complication that requires proper management to prevent the onset of negative outcomes. Patients may experience repeated episodes of cholangitis, secondary biliary cirrhosis, end-stage liver disease and death. OBJECTIVE: To report a single center experience in iatrogenic secondary liver transplantation after cholecystectomy and review the literature. METHODS: This was a retrospective single center study. Of the 1662 liver transplantation realized, 10 (0.60 %) were secondary to iatrogenic bile ducts injuries due cholecystectomies. Medical records of these patients were reviewed in this study. RESULTS: Nine of 10 patients were women; the median time in waiting list and between cholecystectomy and inclusion in waiting list was of 222 days and of 139.9 months, respectively. Cholecystectomy was performed by open approach in eight (80%) cases and by laparoscopic approach in two (20%) cases. The patients underwent an average of 3.5 surgeries and procedures before liver transplantation. Biliary reconstruction was realized with a Roux-en-Y hepaticojejunostomy in nine (90%) cases. Mean operative time was 447.2 minutes and the median red blood cell transfusion was 3.4 units per patient. Mortality in the first month was of 30%. CONCLUSION: Although the liver transplantation is an extreme treatment for an initially benign disease, it has its well-defined indications in treatment of bile duct injuries after cholecystectomy, either in acute or chronic scenario.

RESUMO CONTEXTO: A lesão da via biliar é uma complicação que pode ameaçar a vida e que requer manejo adequado para prevenir o aparecimento de desfechos negativos. Os pacientes podem apresentar episódios repetidos de colangite, cirrose biliar secundária, doença hepática terminal e até mesmo morte. OBJETIVO: Avaliar a experiência de um único centro em transplante hepático secundário a lesão iatrogênica de via biliar pós-colecistectomia e fazer uma revisão de literatura. MÉTODOS: Este foi um estudo retrospectivo de um único centro. Dos 1662 transplantes de fígado, 10 (0,60%) foram secundários a lesões iatrogênicas das vias biliares devido à colecistectomias. Os prontuários médicos desses pacientes foram revisados neste estudo. RESULTADOS: Nove dos dez pacientes eram mulheres; o tempo médio em lista de espera de transplante e entre colecistectomia e inclusão na lista de espera foi de 222 dias e de 139,9 meses, respectivamente. A colecistectomia foi realizada por abordagem aberta em oito (80%) casos e por abordagem laparoscópica em dois (20%) casos. Os pacientes foram submetidos a uma média de 3,5 cirurgias e procedimentos antes do transplante de fígado e a reconstrução biliar foi realizada com hepaticojejunostomia em Y-de-Roux em nove (90%) casos. O tempo operatório médio foi de 447,2 minutos e a média de transfusão de concentrados de hemácias foi de 3,4 unidades por paciente. Mortalidade no primeiro mês foi de 30%. CONCLUSÃO: Embora o transplante de fígado seja um tratamento extremo para uma doença inicialmente benigna, ele tem suas indicações bem definidas no tratamento de lesões biliares após colecistectomia, seja em um cenário agudo ou crônico.

CCAAT/enhancer-binding protein alpha (CEBPA) gene haploinsufficiency does not alter hematopoiesis or induce leukemia in Lck-CALM/AF10 transgenic mice.

Although rare, CALM/AF10 is a chromosomal rearrangement found in immature T-cell acute lymphoblastic leukemia (T-ALL), acute myeloid leukemia, and mixed phenotype acute leukemia of T/myeloid lineages with poor prognosis. Moreover, this translocation is detected in 50% of T-ALL patients with gamma/delta T cell receptor rearrangement, frequently associated with low expression of transcription factor CCAAT/enhancer-binding protein alpha (CEBPA). However, the relevance of CEBPA low expression for CALM/AF10 leukemogenesis has not yet been evaluated. We generated double mutant mice, which express the Lck-CALM/AF10 fusion gene and are haploinsufficient for the Cebpa gene. To characterize the hematopoiesis, we quantified hematopoietic stem cells, myeloid progenitor cells, megakaryocyte-erythrocyte progenitor cells, common myeloid progenitor cells, and granulocyte-macrophage progenitor cells. No significant difference was detected in any of the progenitor subsets. Finally, we tested if Cebpa haploinsufficiency would lead to the expansion of Mac-1+/B220+/c-Kit+ cells proposed as the CALM/AF10 leukemic progenitor. Less than 1% of bone marrow cells expressed Mac-1, B220, and c-Kit with no significant difference between groups. Our results showed that the reduction of Cebpa gene expression in Lck-CALM/AF10 mice did not affect their hematopoiesis or induce leukemia. Our data corroborated previous studies suggesting that the CALM/AF10 leukemia-initiating cells are early progenitors with lymphoid/myeloid differentiating potential.

The Dream of a Mycobacterium.

How do mycobacteria divide? Cell division has been studied extensively in the model rod-shaped bacteria Escherichia coli and Bacillus subtilis, but much less is understood about cell division in mycobacteria, a genus that includes the major human pathogens M. tuberculosis and M. leprae. In general, bacterial cell division requires the concerted effort of many proteins in both space and time to elongate the cell, replicate and segregate the chromosome, and construct and destruct the septum - processes which result in the creation of two new daughter cells. Here, we describe these distinct stages of cell division in B. subtilis and follow with the current knowledge in mycobacteria. As will become apparent, there are many differences between mycobacteria and B. subtilis in terms of both the broad outline of cell division and the molecular details. So, while the fundamental challenge of spatially and temporally organizing cell division is shared between these rod-shaped bacteria, they have solved these challenges in often vastly different ways.

"Among dead and wounded": mapping, characterization and analysis of fires with victims in Recife's metropolitan zone / "Entre mortos e feridos": mapeamento, caracterização e análise dos incêndios com vítimas na região metropolitana do Recife / "Entre muertos y heridos": cartografía, caracterización y análisis de los incendios con víctimas en la zona metropolitana de Recife

This article presents the mapping and analysis of fires with dead and wounded people in the Metropolitan Region of Recife (MRR) served by the Firefighters Department from 2013 to 2016. There was an average rate of 1 death per million inhabitants, similar to countries such as Singapore and Vietnam. The weighted number of fires per wounded or dead person results in rates of 0.5 and 1.7 per 100 recorded fires, respectively. These numbers are concerning, especially when compared to rates from other regions in the world. The victims of fires in MRR were shown to generally not be a perceivable problem in terms of common sense, yet they are real and require accurate analysis and effective measures.

Este artigo apresenta o mapeamento e a análise de incêndios com mortes e feridos na Região Metropolitana do Recife ­ RMR, atendidos pelo Corpo de Bombeiros, no período de 2013 a 2016. Verificou-se uma taxa média de 1 morte por milhão de habitantes, semelhante a países como Singapura e Vietnam. Quando se pondera a quantidade de incêndios para que haja um ferido ou morto, as taxas se apresentam respectivamente em 0,5 e 1,7 por 100 incêndios registrados ­ sendo estes números preocupantes, principalmente quando comparados com taxas de outras regiões no mundo. Conclui-se que as vítimas de incêndios na Região (RMR) são um problema silente ao senso comum, mas real e que exige análise acurada e providências efetivas.

Este artículo presenta la cartografía y análisis de incendios con muertes y heridos en la Zona Metropolitana de Recife (ZMR), atendidos por el Cuerpo de Bomberos, en el período de 2013 a 2016. Se ha verificado un promedio de 1 muerte por millón de habitantes, semejante a países como Singapur y Vietnam. Cuando se examina la cantidad de incendios para que haya un herido o muerto, los promedios se presentan respectivamente de 0,5 y 1,7 por 100 incendios registrados, lo cual es preocupante, principalmente en comparación con los promedios de otras regiones del mundo. Se concluye que las víctimas de incendios en la ZMR son un problema silencioso y real, lo que exige un análisis cuidadoso y diligencias efectivas.

CCAAT/enhancer-binding protein alpha (CEBPA) gene haploinsufficiency does not alter hematopoiesis or induce leukemia in Lck-CALM/AF10 transgenic mice

Although rare, CALM/AF10 is a chromosomal rearrangement found in immature T-cell acute lymphoblastic leukemia (T-ALL), acute myeloid leukemia, and mixed phenotype acute leukemia of T/myeloid lineages with poor prognosis. Moreover, this translocation is detected in 50% of T-ALL patients with gamma/delta T cell receptor rearrangement, frequently associated with low expression of transcription factor CCAAT/enhancer-binding protein alpha (CEBPA). However, the relevance of CEBPA low expression for CALM/AF10 leukemogenesis has not yet been evaluated. We generated double mutant mice, which express the Lck-CALM/AF10 fusion gene and are haploinsufficient for the Cebpa gene. To characterize the hematopoiesis, we quantified hematopoietic stem cells, myeloid progenitor cells, megakaryocyte-erythrocyte progenitor cells, common myeloid progenitor cells, and granulocyte-macrophage progenitor cells. No significant difference was detected in any of the progenitor subsets. Finally, we tested if Cebpa haploinsufficiency would lead to the expansion of Mac-1+/B220+/c-Kit+ cells proposed as the CALM/AF10 leukemic progenitor. Less than 1% of bone marrow cells expressed Mac-1, B220, and c-Kit with no significant difference between groups. Our results showed that the reduction of Cebpa gene expression in Lck-CALM/AF10 mice did not affect their hematopoiesis or induce leukemia. Our data corroborated previous studies suggesting that the CALM/AF10 leukemia-initiating cells are early progenitors with lymphoid/myeloid differentiating potential.

Maturing Mycobacterium smegmatis peptidoglycan requires non-canonical crosslinks to maintain shape.

In most well-studied rod-shaped bacteria, peptidoglycan is primarily crosslinked by penicillin-binding proteins (PBPs). However, in mycobacteria, crosslinks formed by L,D-transpeptidases (LDTs) are highly abundant. To elucidate the role of these unusual crosslinks, we characterized Mycobacterium smegmatis cells lacking all LDTs. We find that crosslinks generate by LDTs are required for rod shape maintenance specifically at sites of aging cell wall, a byproduct of polar elongation. Asymmetric polar growth leads to a non-uniform distribution of these two types of crosslinks in a single cell. Consequently, in the absence of LDT-mediated crosslinks, PBP-catalyzed crosslinks become more important. Because of this, Mycobacterium tuberculosis (Mtb) is more rapidly killed using a combination of drugs capable of PBP- and LDT- inhibition. Thus, knowledge about the spatial and genetic relationship between drug targets can be exploited to more effectively treat this pathogen.