Caracterización de un proteoma mínimo: Mycoplasma genitalium

Comunicaciones a congreso

Comparación de dos técnicas Proteómicas aplicadas al análisis de las proteínas de membrana de Mycoplasma genitalium

Comunicaciones a congreso

Extended proteome and membrane subproteome of mycoplasma genitalium

Comunicaciones a congreso

Legionella SBT applied directly to respiratory samples as a rapid molecular epidemiological tool

Legionnaires' disease (LD) is an atypical pneumonia caused by the inhalation of Legionella. The methods used for the diagnosis of LD are direct culture of respiratory samples and urinary antigen detection. However, the sensitivity of culture is low, and the urinary antigen test is specific only for L. pneumophila sg1. Moreover, as no isolates are obtained, epidemiological studies cannot be performed. The implementation of Nested-sequence-based typing (Nested-SBT) makes it possible to carry out epidemiological studies while also confirming LD, especially in cases caused by non-sg 1. Sixty-two respiratory samples from patients with Legionella clinically confirmed by positive urinary antigen tests were cultured and tested by Nested-SBT, following the European Study Group for Legionella Infections (ESGLI) protocol. Only 2/62 (3.2%) respiratory samples were culture-positive. Amplification and sequencing of Nested-SBT genes were successfully performed in 57/62 samples (91.9%). The seven target genes were characterised in 39/57 (68.4%) respiratory samples, and the complete sequence type (ST) was obtained. The mip gene was the most frequently amplified and sequenced. Nested-SBT is a useful method for epidemiological studies in culture-negative samples, achieving a 28.7-fold improvement over the results of culture studies and reducing the time needed to obtain molecular epidemiological results

Clinical and Epidemiological Characteristics of Streptococcus suis Infections in Catalonia, Spain

Streptococcus suis (S. suis) is a human zoonotic pathogen of occupational origin, with infection acquired through contact with live pigs or pig meat. Pig farming is one of Catalonia's biggest industries and as a result this region of Spain has one of the highest density pig populations per km 2. The aim of our study was to describe the infections caused by S. suis occurring in that area over a 9-year period. A retrospective, multi-center study was carried out by searching records from 15 hospitals in Catalonia for the period between 2010 and 2019. Over the study period altogether nine cases of S. suis infection were identified in five hospitals, with five of these cases occurring in the 2018-2019 period. The mean age of patients was 48 ± 8.9 years and all of them were males. Five patients (55.6%) worked in pig farms. The most frequent manifestation of infection was meningitis (5 cases; 55.6%) followed by septic arthritis (3 cases; 33.3%). None of the patients died at 30 days; nonetheless, 4 developed hearing loss as a long-term complication. The most commonly identified S. suis infection was meningitis. Over 50% of the episodes occurred in the last 2 years and have affected pig farm workers. Further surveillance is needed in order to know its prevalence

Caracterización de un proteoma mínimo: Mycoplasma genitalium

Mycoplasma genitalium es un patógeno humano asociado con algunas enfermedades de transmisión sexual. Las técnicas proteómicas, además de otras técnicas en análisis de la expresión génica, tienen un papel muy importante en el entendimiento de los mecanismo(s) de patogénesis y fisiología bacteriana. El proteoma de Mycoplasma genitalium, modelo de célula mínima, ha sido extendido utilizando diferentes tecnologías y aproximaciones. El proteoma total de este microorganismo ha sido analizado mediante diferentes metodologías que ofrece la proteómica, gracias a las cuales se ha conseguido la identificación del 85.3% de los ORFs predichos. Además, se ha desarrollado el análisis del subproteoma de membrana. En este sentido, la fracción soluble en TX-114 ha sido analizada así como las proteínas de superficie celular utilizando el marcaje de proteínas de superficie mediante CyDye. Finalmente, la respuesta serológica frente a Mycoplasma genitalium de pacientes infectados y de donantes sanos ha sido analizada para conocer que proteínas bacterianas promueven la respuesta inmune. Por consiguiente, no solo se presenta el análisis proteómico más extenso de Mycoplasma genitalium, sino también el análisis proteico más completo de la membrana de Mycoplasma genitalium además de la respuesta serológica en humanos.Mycoplasma genitalium is a human pathogen associated with several sexually transmitted diseases. Proteomic technologies, along with other methods for global gene expression analysis, play a key role in understanding the mechanisms of bacterial pathogenesis and physiology. The proteome of M. genitalium, model of a minimal cell, has been extended using a combination of different proteomic approaches and technologies. The total proteome of this microorganism has been analyzed using gel‐based and gel‐free approaches achieving the identification of 85.3% of the predicted ORFs. In addition, a comprehensive analysis of membrane subproteome has been performed. For this purpose, the TX‐114 soluble fraction has been analyzed as well as the surface proteins, using cell surface protein labeling with CyDye. Finally, the serological response of M. genitalium infected patients and healthy donors has been analyzed to identify proteins that trigger immunological response. Here we present the most extensive M. genitalium proteome analysis (85.3% of predicted ORFs) , a comprehensive M. genitalium membrane analysis, and a study of the human serological response to M. genitalium

Development of an integrated method of concentrationand immunodetection of bacteria

The microbial quality of water is a key aspect to avoid environmental and public health problems. The low pathogen concentration needed to produce a disease outbreak makes it essential to process large water volumes and use sensitive and specific methods such as immunoassays for its detection. In the present work, we describe the development of a device based on microfiltration membranes to integrate the concentration and the immunodetection of waterborne bacteria. A microfiltration membrane treatment protocol was designed to reduce the non-specific binding of antibodies, for which different blocking agents were tested. Thus, the proof of concept of the microbial detection system was also carried out using Escherichia coli as the bacterial pathogen model. E. coli suspensions were filtered through the membranes at 0.5 mL s−1, and the E. coli concentration measurements were made by absorbance, at 620 nm, of the resultant product of the enzymatic reaction among the horseradish peroxidase (HRP) bonded to the antibody, and the substrate 3,3′,5,5′-tetramethylbenzidine (TMB). The results showed that the homemade concentration system together with the developed membrane treatment protocol is able to detect E. coli cells with a limit of detection (LoD) of about 100 CFU in 100 mL.The authors acknowledge financial support from the Spanish Ministry of Economy and Competitiveness (BACSYS pro- ject CTQ2014-54553-C3-1-R). N.U. and O.C. acknowledge funding from the People Programme (Marie Curie Actions) of the 7th Framework Programme of the E uropean Union (FP7/2007-2013), TECNIO-spring program from the Agency for Business Competitiveness of the Government of Catalonia (ACCIÓ). J.J.E. ac- knowledges financial support from the Catalan Industrial Doctorate pro- gram and Waterologies. S.L. and N.P. acknowledge financial support from Gas Natural Fenosa.Peer reviewe

Porta-filtros integrado y procedimiento de concentración y detección de microorganismos

[EN] An integrated filter-holder that enables a target microorganism in a fluid to be concentrated by filtering the fluid and culturing the target mechanism to enable the detection thereof. The filter-holder comprises a top piece (1), an intermediate piece (5) and a bottom piece (4) joined together. A filtration membrane is arranged on the intermediate piece (5) and a reaction chamber (6) is provided over the former for the culture of the microorganism trapped in the filtration membrane when the sample is filtered. The filtration membrane is kept inside the device during the entire concentration and detection method. The invention also relates to the method for the concentration and detection of microorganisms using a filter-holder such as the one described above.[ES] Porta-filtros integrado que permite realizar la concentración de un microorganismo diana de un fluido mediante filtrado de dicho fluido y un cultivo de dicho mecanismo diana para permitir su detección. El porta-filtros comprende una pieza superior (1), una pieza intermedia (5) y una pieza inferior (4) unidas entre sí. En la pieza intermedia (5) se dispone una membrana de filtrado y sobre ésta una cámara de reacción (6)en la que se realiza el cultivo del microorganismo atrapado en la membrana de filtrado al filtrar la muestra. La membrana de filtrado se mantiene en el interior del dispositivo durante todo el procedimiento de concentración y detección. Es también un objeto de la invención el procedimiento de concentración y detección de microorganismos en un porta-filtros como el descrito.Peer reviewedConsejo Superior de Investigaciones Científicas (España), Institut D'Investigació Germans Trias i Pujol, Centro de Investigación Biomédica en Red (CIBER), Universitat Autónoma de BarcelonaA1 Solicitud de patente con informe sobre el estado de la técnic

Porta-filtros integrado y procedimiento de concentración y detección de microorganismos

[EN] An integrated filter-holder that enables a target microorganism in a fluid to be concentrated by filtering the fluid and culturing the target mechanism to enable the detection thereof. The filter-holder comprises a top piece (1), an intermediate piece (5) and a bottom piece (4) joined together. A filtration membrane is arranged on the intermediate piece (5) and a reaction chamber (6) is provided over the former for the culture of the microorganism trapped in the filtration membrane when the sample is filtered. The filtration membrane is kept inside the device during the entire concentration and detection method. The invention also relates to the method for the concentration and detection of microorganisms using a filter-holder such as the one described above.[ES] Porta-filtros integrado que permite realizar la concentración de un microorganismo diana de un fluido mediante filtrado de dicho fluido y un cultivo de dicho mecanismo diana para permitir su detección. El porta-filtros comprende una pieza superior (1), una pieza intermedia (5) y una pieza inferior (4) unidas entre sí. En la pieza intermedia (5) se dispone una membrana de filtrado y sobre ésta una cámara de reacción (6)en la que se realiza el cultivo del microorganismo atrapado en la membrana de filtrado al filtrar la muestra. La membrana de filtrado se mantiene en el interior del dispositivo durante todo el procedimiento de concentración y detección. Es también un objeto de la invención el procedimiento de concentración y detección de microorganismos en un porta-filtros como el descrito.Peer reviewedConsejo Superior de Investigaciones Científicas (España), Institut D'investigació German Trials I Pujol, Centro de Investigación Biomédica en Red (CIBER), Universitat Autònoma de BarcelonaE Solicitud de patente europe

Caracterización de un proteoma mínimo : Mycoplasma genitalium /

Mycoplasma genitalium es un patógeno humano asociado con algunas enfermedades de transmisión sexual. Las técnicas proteómicas, además de otras técnicas en análisis de la expresión génica, tienen un papel muy importante en el entendimiento de los mecanismo(s) de patogénesis y fisiología bacteriana. El proteoma de Mycoplasma genitalium, modelo de célula mínima, ha sido extendido utilizando diferentes tecnologías y aproximaciones. El proteoma total de este microorganismo ha sido analizado mediante diferentes metodologías que ofrece la proteómica, gracias a las cuales se ha conseguido la identificación del 85.3% de los ORFs predichos. Además, se ha desarrollado el análisis del subproteoma de membrana. En este sentido, la fracción soluble en TX-114 ha sido analizada así como las proteínas de superficie celular utilizando el marcaje de proteínas de superficie mediante CyDye. Finalmente, la respuesta serológica frente a Mycoplasma genitalium de pacientes infectados y de donantes sanos ha sido analizada para conocer que proteínas bacterianas promueven la respuesta inmune. Por consiguiente, no solo se presenta el análisis proteómico más extenso de Mycoplasma genitalium, sino también el análisis proteico más completo de la membrana de Mycoplasma genitalium además de la respuesta serológica en humanos.Mycoplasma genitalium is a human pathogen associated with several sexually transmitted diseases. Proteomic technologies, along with other methods for global gene expression analysis, play a key role in understanding the mechanisms of bacterial pathogenesis and physiology. The proteome of M. genitalium, model of a minimal cell, has been extended using a combination of different proteomic approaches and technologies. The total proteome of this microorganism has been analyzed using gel-based and gel-free approaches achieving the identification of 85.3% of the predicted ORFs. In addition, a comprehensive analysis of membrane subproteome has been performed. For this purpose, the TX-114 soluble fraction has been analyzed as well as the surface proteins, using cell surface protein labeling with CyDye. Finally, the serological response of M. genitalium infected patients and healthy donors has been analyzed to identify proteins that trigger immunological response. Here we present the most extensive M. genitalium proteome analysis (85.3% of predicted ORFs) , a comprehensive M. genitalium membrane analysis, and a study of the human serological response to M. genitalium