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Bacterial biofilms form when bacteria attach to surfaces and generate an extracellular matrix that embeds and stabilizes a growing community. Detailed visualization and quantitative analysis of biofilm architecture by optical microscopy are limited by the law of diffraction. Expansion Microscopy (ExM) is a novel Super-Resolution technique where specimens are physically enlarged by a factor of ∼4, prior to observation by conventional fluorescence microscopy. ExM requires homogenization of rigid constituents of biological components by enzymatic digestion. We developed an ExM approach capable of expanding 48-h old Proteus mirabilis biofilms 4.3-fold (termed PmbExM), close to the theoretic maximum expansion factor without gross shape distortions. Our protocol, based on lytic and glycoside-hydrolase enzymatic treatments, degrades rigid components in bacteria and extracellular matrix. Our results prove PmbExM to be a versatile and easy-to-use Super-Resolution approach for enabling studies of P. mirabilis biofilm architecture, assembly, and even intracellular features, such as DNA organization.

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Abstract INTRODUCTION Molecular techniques for the detection of pathogens have been shown to be effective diagnostic tools with high sensitivity and short turnaround times. METHODS This study compared five Staphylococcus aureus DNA extraction methods for detection by the polymerase chain reaction. RESULTS: The concentration and purity of the extracted DNA showed that the methods did not yield DNA of significant quality. However, most protocols yielded 100% positivity, even with low DNA concentrations. CONCLUSIONS: Although one protocol seemed more efficient than the others, PCR was sensitive enough to allow for detection of S. aureus with all the protocols.

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La leptospirosis es una antropozoonosis causada por diferentes serotipos de leptospira cuya epidemiología se asocia con alta precipitación, con una alta incidencia en los animales domésticos, de interés zootécnico y las especies silvestres. Los perros tienen una estrecha relación con los seres humanos y la infección renal puede eliminar las bacterias en la orina, una importante fuente de infección. El presente estudio tuvo como objetivo determinar la prevalencia de anticuerpos y la carga bacteriana de Leptospira spp . en 151 perros recibido en un Centro de Control de Zoonosis. Se recogieron muestras de sangre para obtener suero y la realización de la Prueba de Aglutinación Microscópica (MAT). 59/151 (39,1%) de los perros eran seropositivos a al menos un serovar, con mayor prevalencia y títulos mayores para el serovar Copenhageni (23,73%) del serogrupo icterohaemorrhagiae (16,95%). El serovar canicola, que se encuentra comúnmente en perros, mostró 10,17% de serorreactividad con títulos menores. También se examinaron muestras de suero para detectar la presencia de ADN de la bacteria por la reacción en cadena de la polimerasa convencional (cPCR) utilizando los cebadores LEP1 y LEP2 y sólo una muestra fue positiva, siendo sometida a PCR cuantitativa (qPCR), lo que resultó en un cargo bacterial 14.829.820/mL. Los resultados muestran evidencia de infección con Leptospira spp. en perros, con niveles detectables de anticuerpos MAT y que en un perro podría detectar bacterias en el suero sanguíneo. Es de destacar, por lo tanto, la dispersión de diferente serovares de Leptospira spp. para el medio ambiente, debido a la eliminación de estos serovares por animales portadores renales, así como el riesgo de transmisión a otros animales y seres humanos y la importancia de los perros como animales centinelas en los estudios epidemiológicos.(AU)

Leptospirosis is a anthropozoonosis caused by different serotypes of leptospira whose epidemiology is associated with heavy rainfall, with high occurrence in pets, animals of zootechnical interest and wild species. Dogs have close relationship with humans and in the kidney infection can eliminate the bacteria in the urine, being an important source of infection. The present study aimed to determine the prevalence of antibodies and bacterial load of Leptospira spp. in 151 dogs received at a Center for Zoonosis Control. Blood samples were collected to obtain serum which was submitted to the Microscopic Agglutination Test (MAT). 59/151 (39.1%) dogs were seropositive for at least one serovar with the highest prevalence and higher titers for the serovar Copenhageni (23.73%) serogroup Icterohaemorrhagiae and to sorovar Icterohaemorrhagiae (16.95%). The Canicola serovar, commonly found in dogs, showed 10.17% of seroreactivity with smaller titles. Serum samples were also examined for the presence of DNA of the bacterium by the conventional Polymerase Chain Reaction (cPCR) using the primers Lep1 and Lep2 and only one sample was positive, being subjected to quantitative PCR (qPCR), resulting in bacterial load of 14.829.820/mL. The results show evidence of infection with Leptospira spp. in dogs, with detectable levels of antibodies by MAT, and in one dog the bacteria could be detected in the blood serum. It is noteworthy, therefore, the environment dispersion of different serovars of Leptospira spp., due to the elimination of these serovars by carrier animals kidney, and the risk of transmission to other animals and humans and the importance of dogs as sentinels animals in epidemiological studies.(AU)

A leptospirose é uma antropozoonose causada por diferentes sorotipos de leptospiras cuja epidemiologia está associada a altos índices pluviométricos, com alta ocorrência em animais de companhia, de interesse zootécnico e espécies silvestres. Os cães tem estreita relação com humanos e pela infecção renal podem eliminar a bactéria pela urina, sendo importante fonte de infecção. O presente estudo objetivou a determinação da soroprevalência e a carga bacteriana de Leptospira spp. em 151 cães de um Centro de Controle de Zoonoses. Foram coletadas amostras de sangue para a obtenção de soro e realização da prova de Soroaglutinação Microscópica (SAM). 59/151 (39,1%) dos animais foram sororreagentes para pelo menos um sorovar sendo os de maior prevalência e com títulos mais elevados o sorovar Copenhageni (23,73%) pertencente ao sorogrupo Icterohaemorrhagiae e o sorovar Icterohaemorrhagiae (16,95%). O sorovar Canicola, comumente encontrado nos cães, apresentou 10,17% de sororreatividade com títulos menores. As amostras de soro foram também submetidas a pesquisa de DNA da bactéria pela reação em cadeia pela polimerase convencional (cPCR) utilizando-se os primers Lep1 e Lep2 e apenas uma amostra foi positiva sendo submetida a PCR quantitativa (qPCR), que resultou numa carga bacteriana de 14.829.820/mL. Os resultados demonstram evidência de infecção por Leptospira spp. em cães, com níveis de anticorpos detectáveis à SAM, e que em um cão foi possível detectar a bactéria no soro sanguíneo. Destaca-se, portanto, a dispersão de diferentes sorovares de Leptospira spp. no ambiente, devido à eliminação destes sorovares por animais portadores renais, bem como o risco de transmissão para outros animais e humanos e a importância dos cães como animais sentinelas nos estudos epidemiológicos.(AU)

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Se presenta conceptos sobre la determinación de la composición corporal a través del uso de la técnica de isótopos estables, como el deuterio, mediante el uso de muestras de saliva en escolares de gran altitud. Dichas muestras se encuentran enriquecidas por el óxido de deuterio en todo el cuerpo del escolar y son analizadas con la espectrometría infrarroja con transformada de Fourier. En la actualidad, en el campo de la electromagnética, se hace hincapié sobre una nueva propiedad de las secuencias de ADN bacteriano que inducen a la producción de ondas electromagnéticas, resaltando la memoria que tiene el agua al poder mantener los fragmentos de ADN por largo tiempo, que son generadores de señales, definidos como Biomarcadores, que permitirían la detección de infecciones bacterianas crónicas en el organismo humano a través del uso de la electromagnética. La reflexión a la que se llega, es que muestras de saliva contaminadas con microorganismos, como las bacterias, interfieren con la medición del enriquecimiento de deuterio identificada por el equipo FTIR. Se deja como lección aprendida, el insistir sobre la higiene oral previa a la hora de tomar muestras de saliva, que minimice la contaminación con microorganismos, para lograr determinar el enriquecimiento alcanzado con el deuterio, y así no incurrir en el error de medición que daría resultados no reales del agua corporal total de los sujetos estudiados.

This workpresents concepts about determining body composition byusing the stable isotope technique, as deuterium, using saliva samples in school children at high altitude. These samples are achieved by the enrichment of deuterium oxide in the whole body of the school children and then are analyzed with Fourier transform infrared spectrometer. Currently there is an emphasis on a new property of the bacterial DNA sequences that induce the production of electromagnetic waves, that highlights the water memory to be able to maintain the DNA fragments, signal generators that are defined as Biomarkers, which allow the detection of chronicle bacterial infections in the body. A reflection is that saliva samples contaminated with microorganisms such as bacterias interfere with the measurement of the deuterium enrichment identified by FTIR equipment. The learnt lesson is about oral hygiene, which minimizes contamination with microorganisms from the saliva samples to determine the enrichment achieved with deuterium, and not to make the mistake of measuring actual results would total body water of the subjects studied.

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Immunity against Brucella abortus depends on the recognition of pathogen-associated molecular patterns (PAMPs) by pattern recognition receptors (PRRs). Signaling pathways triggered by Brucella DNA involves TLR9, AIM2 and possibly STING and MAVS. Herein, we review the advances in B. abortus DNA sensing by host innate immune receptors and the progress in this field.

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