Risk assessment for uptake and accumulation of pharmaceuticals by baby leaf lettuce irrigated with reclaimed water under commercial agricultural activities.

The use of reclaimed water to irrigate agricultural crops has increased in recent years as a consequence of water shortage constituting a potential risk for human health. The main objective of this study was to evaluate the impact on the soil-plant system and determining the accumulation of carbamazepine (CBZ), diclofenac (DCF), ketoprofen (KTP) and naproxen (NPX) in the edible part of lettuce under commonly used agricultural practices in commercial production. For this purpose, red oak baby lettuce (Lactuca sativa L.) was irrigated with reclaimed water fortified with different concentrations of pharmaceuticals. The study was carried out in two different scenarios: soil and tray. The tray experiments were conducted with substrate and took place at three different seasons of the year. Lettuce tissue sampled from these experiments were analysed 3 times during the lettuce growing cycle (first, second and third harvest). The practices of first harvest regrowth were also evaluated. For all experiments, CBZ showed the highest accumulation in lettuce leaves of the pharmaceuticals tested, showing a correlation between irrigation exposure time and pharmaceutical uptake. Unexpectedly, DCF was the compound with the highest uptake levels after regrowth practices. Results suggested that pharmaceuticals uptake could be directly associated with the irrigation method and possible accumulation in soil and substrates, while concentration of pharmaceuticals in substrates were 10 times higher. Based on the concentration values detected in lettuce leaves, the risk assessment suggests that no compounds imply any risk to human health, except CBZ for those on vegetarian diets in the tray scenario. Although commercial agricultural practices are usually not considered with regards to risk reduction, in this experiment we demonstrated that climatic conditions are a key factor in pharmaceuticals uptake and different agricultural practices (soil cropping and drip irrigation) can limit the presence of pharmaceutical compounds in crops.

Feral cats do not play a major role in leptospirosis epidemiology on Reunion Island.

Although previous studies have reported Leptospira carriage in kidneys and urine of cats, the role of these animals in leptospirosis epidemiology remains poorly understood. Using molecular methods, we investigated Leptospira renal carriage in 172 feral cats from Reunion Island, an oceanic geographically isolated island located in the South West Indian Ocean. Only one out of the 172 analysed specimens tested positive for Leptospira DNA through quantitative real-time polymerase chain reaction. Using this positive sample, we could obtain sequences at three Leptospira loci (rrs2, lipL32 and lipL41) allowing to report for the first time Leptospira borgpetersenii naturally infecting cats. Comparisons with bacterial sequences from both acute human cases and animal reservoirs revealed similarities with Leptospira sequences previously reported on Reunion Island. However, the low prevalence (0.6%) reported herein does not support any major role of feral cats in leptospirosis epidemiology on Reunion Island, contrasting with results recently reported on another Indian Ocean Island, Christmas Island. The significance of these discrepancies is discussed.

Investigation of a leptospirosis outbreak in triathlon participants, Réunion Island, 2013.

We report herein the investigation of a leptospirosis outbreak occurring in triathlon competitors on Réunion Island, Indian Ocean. All participants were contacted by phone or email and answered a questionnaire. Detection and molecular characterization of pathogenic Leptospira was conducted in inpatients and in rodents trapped at the vicinity of the event. Of the 160 athletes competing, 101 (63·1%) agreed to participate in the study. Leptospirosis was biologically confirmed for 9/10 suspected cases either by real-time PCR or serological tests (MAT or ELISA). The total attack rate, children's attack rate, swimmers' attack rate, and the attack rate in adult swimmers were respectively estimated at 8·1% [95% confidence interval (CI) 4·3-14·7], 0%, 12·7% (95% CI 6·8-22·4) and 23·1% (95% CI 12·6-33·8). Leptospirosis cases reported significantly more wounds [risk ratio (RR) 4·5, 95% CI 1·6-13], wore complete neoprene suits less often (RR 4·3, 95% CI 1·3-14·5) and were most frequently unlicensed (RR 6·6, 95% CI 2·9-14·8). The epidemiological investigation supported that some measures such as the use of neoprene suits proved efficient in protecting swimmers against infection. PCR detection in rats revealed high Leptospira infection rates. Partial sequencing of the 16S gene and serology on both human and animal samples strongly suggests that rats were the main contaminators and were likely at the origin of the infection in humans.

Chikungunya-Wolbachia interplay in Aedes albopictus.

A severe Chikungunya (CHIK) outbreak recently hit several countries of the Indian Ocean. On La Réunion Island, Aedes albopictus was incriminated as the major vector. This mosquito species is naturally co-infected with two distinct strains of the endosymbiont Wolbachia, namely wAlbA and wAlbB, which are increasingly attracting interest as potential tools for vector control. A PCR quantitative assay was developed to investigate Wolbachia/mosquito host interactions. We show that Wolbachia densities are slightly decreased in CHIK virus (CHIKV)-infected females. We measured the impact of CHIKV replication on a lysogenic virus: WO bacteriophage. Our data indicate that WO is sheltered by wAlbB, likely at a single copy per bacteria, and that CHIKV replication is not a physiological stress triggering WO entrance into the lytic cycle.

Plantilla de la sonrisa / Smiles’s Template

La valoración estética en reposo y en sonrisa es fundamental en la exploración clínica del paciente ortodóncico, dentro de la cual el análisis y diseño de la sonrisa se ha convertido en la última década en la llave del diagnóstico y del tratamiento. Se ha realizado una amplia revisión bibliográfica, donde los autores coinciden en valorar la sonrisa por la exposición gingival, por su relación labiodental, simetría y tamaño transversal de forma individual; sin embargo, de momento no existe en la bibliografía un modo de evaluar cada uno de estos factores de manera integral. Con el fin de incorporar el análisis exhaustivo de la sonrisa en la práctica clínica diaria, resulta necesario el uso de una plantilla a modo de esquema que nos permita registrar de manera rápida y sistemática cada una de las variables

The aesthetic evaluation of the face, both relaxed and smiling, is basic in the clinical exploration of the orthodontic patient. Within this valuation, the analysis and the design of the smile, has become the most important key issue for both the diagnostic and the treatment in the last decade. Extensive bibliographic revision has been made through which different authors agree to analyze the smile by its gingival exposure, lip and teeth relationship, symmetry, and transversal measure among others. Individually you cannot find throughout this literature a way to measure each of this factors in an integral way. In order to incorporate the exhaustive analysis of the smile on a daily clinical practice, it becomes necessary the use of a template as a scheme that allows us to quickly and systematically register each of these variables

[Intrapleural urokinase in the treatment of parapneumonic effusions]. / Utilidad de la urocinasa intrapleural en el tratamiento del derrame pleural paraneumónico.

INTRODUCTION: Intrapleural fibrinolytic instillation has been used in the treatment of loculated pleural effusions and empyemas and has reduced the need for surgical intervention. Currently, the most commonly used fibrinolytic is urokinase, although the doses have not yet been standardized in children. The aim of the present study was to evaluate the utility of urokinase in the treatment of infectious pleural effusions in children. MATERIAL AND METHODS: A retrospective study was performed of children with infectious pleural effusions admitted to the pediatric intensive care unit (PICU) between January 2000 and December 2003. Age, sex, clinical features, laboratory tests, response to urokinase treatment and clinical course during hospital stay were analyzed. RESULTS: Thirty-one children were treated. The mean age was 38.1 months (SD: 22). There were 18 boys and 13 girls. The most frequent month of diagnosis was November and the number of admission significantly increased from 2002 onwards. The most frequent antibiotic therapy used before admission to the PICU was cefotaxime associated with vancomycin (41 %), followed by cefotaxime alone (16 %). Positive cultures for Streptococcus pneumoniae were found in 11 patients (35 %). Pleural loculation was found in 14 patients (45 %). Treatment with intrapleural urokinase was used in 23 patients (74 %). The mean chest tube drainage was 140 ml (SD: 175) in the 24 hours before urokinase instillation and was 406 ml (SD: 289) in the 48 hours after fibrinolytic therapy (p < 0.05). Twenty-one patients (91 %) who received urokinase treatment had a good response. There were no complications during the treatment. The mean length of stay in the PICU was 5.8 days (SD: 2.6). CONCLUSIONS: The incidence of complicated pleural effusions due to S. pneumoniae has increased in the last few years, despite antibiotic therapy. Intrapleural urokinase is an effective treatment, including in empyemas without loculation. None of our patients required thoracotomy and there were few adverse effects.

Utilidad de la urocinasa intrapleural en el tratamiento del derrame pleural paraneumónico / Intrapleural urokinase in the treatment of parapneumonic effusions

Introducción: La instilación intrapleural de agentes fibrinolíticos se ha utilizado como tratamiento de derrames pleurales tabicados y de empiemas, disminuyendo la necesidad de intervención quirúrgica. La urocinasa es el fibrinolítico más utilizado aunque en pediatría las dosis no están claramente estandarizadas. El objetivo del estudio fue evaluar la utilidad de la urocinasa intrapleural como tratamiento de los derrames pleurales complicados. Material y métodos: Se realizó un estudio retrospectivo de los derrames pleurales ingresados en la unidad de cuidados intensivos pediátricos (UCIP) entre enero de 2000 y diciembre de 2003. Se recogieron las variables de edad, sexo, sintomatología, pruebas complementarias, respuesta al tratamiento con urocinasa y evolución durante su ingreso. Resultados: Se estudiaron 31 casos, la edad media fue de 38,1 meses (desviación estándar [DE]: 22); 18 varones y 13 mujeres. El mes de mayor incidencia fue noviembre, y existió un aumento significativo del número de ingresos a partir del año 2002. El tratamiento antibiótico más frecuente antes del ingreso en UCIP fue la asociación de cefotaxima y vancomicina (41 %), seguido de cefotaxima (16 %). Se obtuvieron cultivos positivos a S. pneumoniae en 11 casos (35 %). Se demostró derrame tabicado en 14 casos (45 %). El tratamiento con urocinasa intrapleural se realizó en 23 casos (74 %), observándose un aumento significativo del drenaje pleural tras su aplicación, con un drenaje medio 24 h previo a la urocinasa de 140 ml (DE: 175) frente a 406 ml (DE: 289) 48 h después (p < 0,05). Respondieron favorablemente al tratamiento 21 casos (91 %) y no existieron complicaciones importantes durante su administración. La duración media del drenaje torácico fue de 5,2 días (DE: 2,97). La estancia media en la UCIP fue de 5,8 días (DE: 2,6). Conclusiones: Existe una mayor incidencia en los últimos años de derrames pleurales complicados secundarios a S. pneumoniae a pesar del tratamiento antibiótico. La urocinasa intrapleural es un tratamiento efectivo incluso en empiemas no tabicados, no precisando en ningún caso la intervención quirúrgica y con escasos efectos secundarios

Introduction: Intrapleural fibrinolytic instillation has been used in the treatment of loculated pleural effusions and empyemas and has reduced the need for surgical intervention. Currently, the most commonly used fibrinolytic is urokinase, although the doses have not yet been standardized in children. The aim of the present study was to evaluate the utility of urokinase in the treatment of infectious pleural effusions in children. Material and methods: A retrospective study was performed of children with infectious pleural effusions admitted to the pediatric intensive care unit (PICU) between January 2000 and December 2003. Age, sex, clinical features, laboratory tests, response to urokinase treatment and clinical course during hospital stay were analyzed. Results: Thirty-one children were treated. The mean age was 38.1 months (SD: 22). There were 18 boys and 13 girls. The most frequent month of diagnosis was November and the number of admission significantly increased from 2002 onwards. The most frequent antibiotic therapy used before admission to the PICU was cefotaxime associated with vancomycin (41 %), followed by cefotaxime alone (16 %). Positive cultures for Streptococcus pneumoniae were found in 11 patients (35 %). Pleural loculation was found in 14 patients (45 %). Treatment with intrapleural urokinase was used in 23 patients (74 %). The mean chest tube drainage was 140 ml (SD: 175) in the 24 hours before urokinase instillation and was 406 ml (SD: 289) in the 48 hours after fibrinolytic therapy (p < 0.05). Twenty-one patients (91 %) who received urokinase treatment had a good response. There were no complications during the treatment. The mean length of stay in the PICU was 5.8 days (SD: 2.6). Conclusions: The incidence of complicated pleural effusions due to S. pneumoniae has increased in the last few years, despite antibiotic therapy. Intrapleural urokinase is an effective treatment, including in empyemas without loculation. None of our patients required thoracotomy and there were few adverse effects

Sites of positive and negative regulation in the Bacillus subtilis antiterminators LicT and SacY.

The Bacillus subtilis homologous transcriptional antiterminators LicT and SacY control the inducible expression of genes involved in aryl beta-glucoside and sucrose utilization respectively. Their RNA-binding activity is carried by the N-terminal domain (CAT), and is regulated by two similar C-terminal domains (PRD1 and PRD2), which are the targets of phosphorylation reactions catalysed by the phosphoenolpyruvate: sugar phosphotransferase system (PTS). In the absence of the corresponding inducer, LicT is inactivated by BglP, the PTS permease (EII) specific for aryl beta-glucosides, and SacY by SacX, a negative regulator homologous to the EII specific for sucrose. LicT, but not SacY, is also subject to a positive control by the general PTS components EI and HPr, which are thought to phosphorylate LicT in the absence of carbon catabolite repression. Construction of SacY/LicT hybrids and mutational analysis enabled the location of the sites of this positive regulation at the two phosphorylatable His207 and His269 within LicT-PRD2, and suggested that the presence of negative charges at these sites is sufficient for LicT activation in vivo. The BglP-mediated inhibition process was found to essentially involve His100 of LicT-PRD1, with His159 of the same domain playing a minor role in this regulation. In vitro experiments indicated that His100 could be phosphorylated directly by the general PTS proteins, this phosphorylation being stimulated by phosphorylated BglP. We confirmed that, similarly, the corresponding conserved His99 residue in SacY is the major site of the negative control exerted by SacX on SacY activity. Thus, for both antiterminators, the EII-mediated inhibition process seems to rely primarily on the presence of a negative charge at the first conserved histidine of the PRD1.

Specificity and genetic polymorphism of the Bacillus competence quorum-sensing system.

A quorum-sensing mechanism involving the pheromone ComX and the ComP-ComA two-component system controls natural competence in Bacillus subtilis. ComX is expressed as a cytoplasmic inactive precursor that is released into the extracellular medium as a cleaved, modified decapeptide. This process requires the product of comQ. In the presence of ComX, the membrane-localized ComP histidine kinase activates the response regulator ComA. We compared the sequences of the quorum-sensing genes from four closely related bacilli, and we report extensive genetic polymorphism extending through comQ, comX, and the 5' two-thirds of comP. This part of ComP encodes the membrane-localized and linker domains of the sensor protein. We also determined the sequences of the comX genes of four additional wild-type bacilli and tested the in vivo activities of all eight pheromones on isogenic strains containing four different ComP receptor proteins. A striking pattern of specificity was discovered, providing strong evidence that the pheromone contacts ComP directly. Furthermore, we show that coexpression of comQ and comX in Escherichia coli leads to the production of active pheromone in the medium, demonstrating that comQ is the only dedicated protein required for the processing, modification, and release of active competence pheromone. Some of the implications of these findings for the evolution and the mechanism of the quorum-sensing system are discussed.

Characterization of ylbF, a new gene involved in competence development and sporulation in Bacillus subtilis.

We used mini Tn10 transposition to generate a library of Bacillus subtilis insertion mutants, with the goal of identifying and characterizing new competence genes. Two new regulatory genes were identified in our screen: ypuN (also known as rsiX, the anti-sigmaX factor) and ylbF. The disruption of ylbF leads to a dramatic decrease in the expression of comK, encoding the competence transcription factor. Our data show that ylbF positively controls ComK at a post-transcriptional level. It has been reported previously that ComK is degraded in vivo and in vitro by a multimeric protein complex composed of ClpP, ClpC and MecA. This proteolysis is inhibited by the ComS peptide. We show that both the overexpression of comS and the inactivation of mecA individually suffice to bypass the competence phenotype of the ylbF mutation. This mutation does not seem to alter the cellular concentrations of MecA or ClpP, and we propose a role for YlbF in modulating the translation, stability or activity of ComS. In addition to its role in competence, ylbF also appears to regulate sporulation by acting before stage II.

Competence for transformation: a matter of taste.

Genetic competence in both Bacillus subtilis and Streptococcus pneumoniae, as well as virulence in Staphylococcus aureus, are regulated by quorum-sensing mechanisms that use two-component signal transduction systems to respond to extracellular peptide pheromones. Recent data indicate that in all three systems closely related strains express markedly different pheromones and polytopic membrane receptor proteins. This polymorphism may function as a sexual isolation mechanism. In B. subtilis the downstream segment of the competence regulatory pathway acts by controlling the stability of a key transcription factor. In S. pneumoniae the downstream segment involves the transcriptional activation of a minor sigma factor that is in turn responsible for the expression of late competence genes.

Mutational analysis and membrane topology of ComP, a quorum-sensing histidine kinase of Bacillus subtilis controlling competence development.

ComP is a sensor histidine kinase of Bacillus subtilis required for the signal transduction pathway that initiates the development of competence for genetic transformation. It is believed that ComP senses the presence of ComX, a modified extracellular peptide pheromone, and donates a phosphate to ComA, thereby activating this transcription factor for binding to the srfA promoter. In the present study, fusions to the Escherichia coli proteins PhoA and LacZ and analysis of its susceptibility to the protease kallikrein were used to probe the membrane topology of ComP. These data suggest that ComP contains six or eight membrane-spanning segments and two large extracytoplasmic loops in its N-terminal membrane-associated domain. Deletions were introduced involving the large extracellular loops to explore the role of the N-terminal domain of ComP in signal transduction. The absence of the second loop conferred a phenotype in which ComP was active in the absence of ComX. The implications of these data are discussed.

Multiple phosphorylation of SacY, a Bacillus subtilis transcriptional antiterminator negatively controlled by the phosphotransferase system.

The Bacillus subtilis SacY transcriptional antiterminator is a regulator involved in sucrose-promoted induction of the sacB gene. SacY activity is negatively controlled by enzyme I and HPr, the general energy coupling proteins of the phosphoenolpyruvate:sugar phosphotransferase system (PTS), and by SacX, a membranal protein homologous to SacP, the B. subtilis sucrose-specific PTS-permease. Previous studies suggested that the negative control exerted by the PTS on bacterial antiterminators of the SacY family involves phosphoenolpyruvate-dependent phosphorylation by the sugar-specific PTS-permeases. However, data reported herein show direct phosphorylation of SacY by HPr(His approximately P) with no requirement for SacX. Experiments were carried out to determine the phosphorylatable residues in SacY. In silico analyses of SacY and its homologues revealed the modular structure of these proteins as well as four conserved histidines within two homologous domains (here designated P1 and P2), present in 14 distinct mRNA- and DNA-binding bacterial transcriptional regulators. Single or multiple substitutions of these histidyl residues were introduced in SacY by site-directed mutagenesis, and their effects on phosphorylation and antitermination activity were examined. In vitro phosphorylation experiments showed that SacY was phosphorylated on three of the conserved histidines. Nevertheless, in vivo studies using cells bearing a sacB'-lacZ reporter fusion, as well as SacY mutants lacking the phosphorylatable histidyls, revealed that only His-99 is directly involved in regulation of SacY antitermination activity.

Analysis of HLA haplotypes in families with type 1 diabetes mellitus in La Réunion island.

To analyse HLA and insulin-dependent diabetes mellitus (IDDM) association in the ethnically mixed population of La Réunion island, we carried out a family study on 70 diabetic subjects. HLA-DQA1, -DQB1 and -DRB1 typing was performed by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP), completed by PCR-sequence-specific oligonucleotide (SSO) and PCR-sequence-specific priming (SSP). Haplotype-relative risks (HRR) were determined with the non-transmitted parental haplotypes as controls, and relative risks (RR) were calculated with a classical case-control study. The most significant risks were found for the cis and trans combinations between DQA1*03 or *0501 (Arg52+) and DQB1*02 or *0302 (Asp57-) alleles, suggesting a direct role for the HLA-DQ heterodimer in IDDM susceptibility. Interestingly, due to the mixed origin of the population, the trans-encoded DQ molecules in the (DR3)-DQA1*0501-DQB1*02/(DR4)-DQA1*03-DQB1*0302 subjects were also found cis-encoded in patients with the (DR7 or 9)-DQA1*03-DQB1*02 haplotype and in a patient with the rare (DR11)-DQA1*0501-DQB1*0302 haplotype. A relative predispositional effect (RPE) analysis gave significant haplotype-IDDM+ associations in the following order: (DR3)-DQA1*0501-DQB1*02 > (DR4)-DQA1*03-DQB1*0302 > (DR9)-DQA1*03- DQB*02 > (DR7)-DQA1*03-DQB1*02 > (DR2)-DQA1*01-DQB1*0502. No protective effect remained significant once the susceptible haplotypes were removed. A stratification study showed a stronger influence of the DQ genes than DRB1 alleles within the DR7 haplotypes. On the other hand, IDDM subjects with only one susceptible haplotype had inherited this haplotype more often from their father than from their mother. This paternal effect could be related to the greater risk of IDDM in offspring of diabetic fathers than the risk in offspring of diabetic mothers.

A ribonucleic antiterminator sequence (RAT) and a distant palindrome are both involved in sucrose induction of the Bacillus subtilis sacXY regulatory operon.

The Bacillus subtilis sacXY regulatory operon is involved in sucrose induction of the levansucrase sacB gene by an antitermination mechanism. In the presence of sucrose, the activated SacY antiterminator protein stabilizes the secondary structure of a ribonucleic antiterminator sequence (RAT) located in the leader region of the sacB transcript, and overlapping a rho-independent transcription terminator. Formation of the SacY-RAT complex prevents alternative formation of the terminator, allowing transcription of the downstream sequences. In the absence of sucrose, inhibition of SacY activity by SacX leads to termination of transcription. Expression of sacXY is also sucrose-inducible. This induction was previously shown to be mediated by SacY itself and/or SacT, another antiterminator involved in induction of genes belonging to a distinct sucrose pathway. These antiterminators are not activated at the same concentration of sucrose. We show here that sacXY induction occurs through activation of either SacY or SacT antiterminators, at their respective sucrose activation concentration. This result demonstrates a link between SacY- and SacT-mediated metabolic pathways. In addition, the sacXY leader region carries a RAT-like sequence, which however does not appear to overlap any apparent rho-independent transcription terminator. Site-directed mutagenesis experiments on this RAT-like sequence demonstrated its involvement in sucrose induction. Deletions generated in the sacXY leader region showed that a palindrome, located 100 nt downstream from the RAT-like sequence, also acts as a cis-acting element. Computer analysis of the leader RNA suggested that formation of the secondary structure of the RAT-like sequence and the palindrome could be mutually exclusive.

[Non-immune hydrops fetalis. Review of 11 cases]. / Hydrops fetalis no inmune. Revisión de 11 casos.

Eleven cases of non-immune hydrops fetalis (NIHF) are presented. Incidence was 1.8 out of 10,000 births. NIHF became more common than immune hydrops fetalis (1.4/10,000 births). Mortality was of 81% and complications were frequent. NIHF is associated with prematurity, neonatal anoxia, polyhydramnios, hypoalbuminemia and hypoproteinemia. Considerable emphasis must be placed on antenatal diagnosis to achieve a precocious treatment and so improve the present poor prognosis.

[Botulism in the infant. Presentation of a case]. / Botulismo del lactante. Presentación de un caso.

First case reported in our country with infant botulism is described in a 5 month old infant. Clinical and electrodiagnostic abnormalities are the only relatively specific findings in infant botulism, which were seen in this infant, but it requires isolation of "C. botulinum" for diagnosis confirmation. "C. botulinum" type B spores were identified in stool samples from this infant. It has been described frequently isolation of "C. botulinum" from honey specimens that had been fed to infants who subsequently developed infant botulism. They are now studying honey samples with appropriate methods (MDL-10), obtained from alimentation of this infant.

[Advances in the treatment of endotoxic shock]. / Avances en el tratamiento del "shock" endotóxico.

Authors present their adopted measures, in the different clinical stages of endotoxic shock with a phisiological approach, paying special attention to those secondary to meningococcal infections.