A DNA pooling based system to detect Escherichia coli virulence factors in fecal and wastewater samples

The availability of a useful tool for simple and timely detection of the most important virulent varieties of Escherichia coli is indispensable. To this end, bacterial DNA pools which had previously been categorized were obtained from isolated colonies as well as selected in terms of utilized phenotype; the pools were assessed by two PCR Multiplex for the detection of virulent E. coli eaeA, bfpA, stx1, stx2, ipaH, ST, LT, and aatA genes, with the 16S gene used as DNA control. The system was validated with 66 fecal samples and 44 wastewater samples. At least one positive isolate was detected by a virulent gene among the 20 that were screened. The analysis of fecal samples from children younger than 6 years of age detected frequencies of 25% LT positive strains, 8.3% eae, 8.3% bfpA, 16.7% ipaH, as well as 12.5 % aatA and ST. On the other hand, wastewater samples revealed frequencies of 25.7% eaeA positive, 30.3% stx1, 15.1% LT and 19.7% aatA. This study is an initial step toward carrying out epidemiological field research that will reveal the presence of these bacterial varieties.UCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias de la Salud::Instituto de Investigaciones en Salud (INISA)UCR::Vicerrectoría de Docencia::Salud::Facultad de Microbiologí

Relation between coliphages presence in drinking water, rain and acute diarrhea in Costa Rica

La calidad del agua de consumo es una de las preocupaciones más grandes a las que se enfrentan los entes de salud. En Costa Rica, a pesar de la buena calidad de las fuentes de agua y del agua de consumo, se siguen registrando numerosos casos de diarrea anualmente. En este estudio se realizó un análisis de indicadores de contaminación virales y bacterianos, en dos comunidades del Valle Central con acueducto propio. Se analizaron en total 24 muestras de agua por cada comunidad, durante un año, 12 en la fuente de agua y 12 en el agua tratada. La frecuencia de diarreas en cada comunidad fue comparada con la calidad microbiológica del agua consumida, encontrándose ausencia de indicadores bacterianos en muestras de agua tratada, mientras que la presencia de indicadores virales en el agua de consumo (con cloro residual) se relaciona con un incremento en el número de casos de diarreas, adicionalmente se analizó el comportamiento de las diarreas de acuerdo a la precipitación promedio con el fin de observar posibles patrones de estacionalidad. Estos hallazgos respaldan la necesidad de ampliar el monitoreo de calidad microbiológica del agua con otro tipo de indicadores de contaminación que correlacionen mejor con otros agentes patógenos de transmisión hídrica, principalmente en época seca, periodo en que se observan más casos de diarrea.The quality of potable water is one of the most important issues of public health. In Costa Rica, in spite of having good sources of water and good quality of water for consumption, there continues to be a significant number of diarrhea cases annually. In this investigation, viral and bacterial analyses of indicators of contamination were conducted in two rural communities with their own aqueducts in the Central Valley of Costa Rica. Twenty - four samples were taken from each community during the year; 12 fro m the water sources and 12 from the treated waters. The frequency of diarrhea in each community was compared to the microbiological quality of the water being consumed, noting an absence of bacterial indicators in samples of treated water, while the presen ce of viral indicators in the water (with residual chlorine) were associated with an incremental increase in the number of cases of diarrhea. The analysis also included measurements of average monthly rainfall with the aim of observing possible association s between seasonal conditions and the incidence of diarrhea cases. These findings indicate the need to increase the monitoring of the microbiological quality of the water with other types of indicators of contamination that correlate better with other path ogens transmitted by water, especially in the dry seasons when a greater number of cases of diarrhea are observed.UCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias de la Salud::Instituto de Investigaciones en Salud (INISA

Seven-versus 14-day course of antibiotics for the treatment of bloodstream infections by Enterobacterales: a randomized, controlled trial

[Objective] To prove that 7-day courses of antibiotics for bloodstream infections caused by members of the Enterobacterales (eBSIs) allow a reduction in patients' exposure to antibiotics while achieving clinical outcomes similar to those of 14-day schemes.[Methods] A randomized trial was performed. Adult patients developing eBSI with appropriate source control were assigned to 7 or 14 days of treatment, and followed 28 days after treatment cessation; treatments could be resumed whenever necessary. The primary endpoint was days of treatment at the end of follow-up. Clinical outcomes included clinical cure, relapse of eBSI and relapse of fever. A superiority margin of 3 days was set for the primary endpoint, and a non-inferiority margin of 10% was set for clinical outcomes. Efficacy and safety were assessed together with a DOOR/RADAR (desirability of outcome ranking and response adjusted for duration of antibiotic risk) analysis.[Results] 248 patients were assigned to 7 (n = 119) or 14 (n = 129) days of treatment. In the intention-to-treat analysis, median days of treatment at the end of follow-up were 7 and 14 days (difference 7, 95%CI 7–7). The non-inferiority margin was also met for clinical outcomes, except for relapse of fever (–0.2%, 95%CI –10.4 to 10.1). The DOOR/RADAR showed that 7-day schemes had a 77.7% probability of achieving better results than 14-day treatments.[Conclusions] 7-day schemes allowed a reduction in antibiotic exposure of patients with eBSI while achieving outcomes similar to those of 14-day schemes. The possibility of relapsing fever in a limited number of patients, without relevance to final outcomes, may not be excluded, but was overcome by the benefits of shortening treatments.This work received technical support from the Spanish Network of Research in Infectious Diseases (REIPI) and the Spanish Clinical Research Network (SCReN) of the Instituto de Salud Carlos III, Ministerio de Economía y Competitividad, Spanish Government (PT13/0002/0010).Peer reviewe

Effectiveness of Fosfomycin for the Treatment of Multidrug-Resistant Escherichia coli Bacteremic Urinary Tract Infections

IMPORTANCE The consumption of broad-spectrum drugs has increased as a consequence of the spread of multidrug-resistant (MDR) Escherichia coli. Finding alternatives for these infections is critical, for which some neglected drugs may be an option. OBJECTIVE To determine whether fosfomycin is noninferior to ceftriaxone or meropenem in the targeted treatment of bacteremic urinary tract infections (bUTIs) due to MDR E coli. DESIGN, SETTING, AND PARTICIPANTS This multicenter, randomized, pragmatic, open clinical trial was conducted at 22 Spanish hospitals from June 2014 to December 2018. Eligible participants were adult patients with bacteremic urinary tract infections due to MDR E coli; 161 of 1578 screened patients were randomized and followed up for 60 days. Data were analyzed in May 2021. INTERVENTIONS Patients were randomized 1 to 1 to receive intravenous fosfomycin disodium at 4 g every 6 hours (70 participants) or a comparator (ceftriaxone or meropenem if resistant; 73 participants) with the option to switch to oral fosfomycin trometamol for the fosfomycin group or an active oral drug or pa renteral ertapenem for the comparator group after 4 days. MAIN OUTCOMES AND MEASURES The primary outcome was clinical and microbiological cure (CMC) 5 to 7 days after finalization of treatment; a noninferiority margin of 7% was considered. RESULTS Among 143 patients in the modified intention-to-treat population (median [IQR] age, 72 [62-81] years; 73 [51.0%] women), 48 of 70 patients (68.6%) treated with fosfomycin and 57 of 73 patients (78.1%) treated with comparators reached CMC (risk difference, -9.4 percentage points; 1-sided 95% CI, -21.5 to infinity percentage points; P = .10). While clinical or microbiological failure occurred among 10 patients (14.3%) treated with fosfomycin and 14 patients (19.7%) treated with comparators (risk difference, -5.4 percentage points; 1-sided 95% CI. -infinity to 4.9; percentage points; P = .19), an increased rate of adverse event-related discontinuations occurred with fosfomycin vs comparators (6 discontinuations [8.5%] vs 0 discontinuations; P = .006). In an exploratory analysis among a subset of 38 patients who underwent rectal colonization studies, patients treated with fosfomycin acquired a new ceftriaxone-resistant or meropenem-resistant gram-negative bacteria at a decreased rate compared with patients treated with comparators (0 of 21 patients vs 4 of 17 patients [23.5%]; 1-sided P = .01). CONCLUSIONS AND RELEVANCE This study found that fosfomycin did not demonstrate noninferiority to comparators as targeted treatment of bUTI from MDR E coli; this was due to an increased rate of adverse event-related discontinuations. This finding suggests that fosfomycin may be considered for selected patients with these infections

Treatment with tocilizumab or corticosteroids for COVID-19 patients with hyperinflammatory state: a multicentre cohort study (SAM-COVID-19)

Objectives: The objective of this study was to estimate the association between tocilizumab or corticosteroids and the risk of intubation or death in patients with coronavirus disease 19 (COVID-19) with a hyperinflammatory state according to clinical and laboratory parameters. Methods: A cohort study was performed in 60 Spanish hospitals including 778 patients with COVID-19 and clinical and laboratory data indicative of a hyperinflammatory state. Treatment was mainly with tocilizumab, an intermediate-high dose of corticosteroids (IHDC), a pulse dose of corticosteroids (PDC), combination therapy, or no treatment. Primary outcome was intubation or death; follow-up was 21 days. Propensity score-adjusted estimations using Cox regression (logistic regression if needed) were calculated. Propensity scores were used as confounders, matching variables and for the inverse probability of treatment weights (IPTWs). Results: In all, 88, 117, 78 and 151 patients treated with tocilizumab, IHDC, PDC, and combination therapy, respectively, were compared with 344 untreated patients. The primary endpoint occurred in 10 (11.4%), 27 (23.1%), 12 (15.4%), 40 (25.6%) and 69 (21.1%), respectively. The IPTW-based hazard ratios (odds ratio for combination therapy) for the primary endpoint were 0.32 (95%CI 0.22-0.47; p < 0.001) for tocilizumab, 0.82 (0.71-1.30; p 0.82) for IHDC, 0.61 (0.43-0.86; p 0.006) for PDC, and 1.17 (0.86-1.58; p 0.30) for combination therapy. Other applications of the propensity score provided similar results, but were not significant for PDC. Tocilizumab was also associated with lower hazard of death alone in IPTW analysis (0.07; 0.02-0.17; p < 0.001). Conclusions: Tocilizumab might be useful in COVID-19 patients with a hyperinflammatory state and should be prioritized for randomized trials in this situatio

Wastewater-based epidemiology of enteric viruses and surveillance of acute gastrointestinal illness outbreaks in a resource-limited region

Acute gastrointestinal illness (AGI) continues to be an important cause of morbidity and mortality among all ages. This study applied the principles ofwastewater-based epidemiology for the preventive identification of potential outbreaks of acute viral gastroenteritis and hepatitis A by analyzing the presence of human enteric viruses in influents of small municipal wastewater treatment plants (WWTPs) handling domestic sewage, together with public health reports of acute diarrheal and hepatitis A disease in Costa Rica during 2013. Raw wastewater samples were collected during four seasonal periods with different rainfall levels. The presence of five human enteric viruses (rotavirus A, norovirus GI, norovirus GII, enterovirus, and hepatitis A virus) was studied by endpoint and real-time polymerase chain reaction in influents of five WWTPs. Cases of AGI were analyzed using historical public health reports of endemic levels and quartile ranges for each administrative and territorial area where theWWTPs are located and for its surrounding counties. A tendency for a higher rate of positive viral tests was present 1 week before an increase of AGI cases. Epidemiological weeks categorized asOutbreak (above the 75th percentile) and Success (below the 25th percentile) showed statistically significant differences in terms of positive viral test rates (Wilcoxon test, P50.05). Virologicalmonitoring ofwastewater in smallWWTPs is an appropriate model for epidemiological surveillance of diarrheal and hepatitis A diseases in low- and middle-resource countries.Universidad de Costa Rica/[]/UCR/Costa RicaInstituto Costarricense de Acueductos y Alcantarillados/[]/AyA/Costa RicaUCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias de la Salud::Instituto de Investigaciones en Salud (INISA

Remotion of physicochemical and microbiological parameters on wastewater treatment plants in Costa Rica’s Great Metropolitan Area

Las aguas residuales inadecuadamente tratadas son un factor de riesgo para la salud pública dado que son un vehículo de diseminación de microorganismos patógenos como virus entéricos. Sin embargo, la circulación de patógenos virales en esta matriz ha sido poco estudiada en la región. El objetivo de este trabajo fue describir la presencia de cinco virus enteropatógenos y parámetros operativos básicos, en afluentes y efluentes de cinco plantas de tratamiento de aguas residuales (PTAR) con sistemas de lodos activados, ubicadas en el Gran Área Metropolitana de Costa Rica durante el 2013. Los patógenos virales estudiados fueron: enterovirus, virus de hepatitis A, rotavirus A, norovirus G1 y norovirus G2 (determinados por PCR punto final); mientras que los parámetros de control de rendimiento analizados incluyeron disponibilidad bioquímica de oxígeno, disponibilidad química de oxígeno, sólidos suspendidos, sólidos sedimentables, grasas y aceites, sustancias activas al azul de metileno y coliformes fecales. A nivel fisicoquímico todas las plantas mostraron un rendimiento aceptable, caso contrario a los parámetros microbiológicos, todos los virus investigados fueron detectados en aguas tratadas y no tratadas durante todo el año, siendo más frecuentes durante la época seca. El virus más frecuente fue rotavirus, seguido por norovirus G1. En conclusión, este estudio describe que durante el periodo estudiado las PTAR fueron eficientes en la remoción de parámetros fisicoquímicos, y que la circulación de virus entéricos es común en las PTAR estudiadas. Por ello, es necesaria la inclusión de procesos de desinfección adicionales para una descarga segura de las aguas tratadas.Inefficient treated wastewaters are a critical factor for public health due to can be a dissemination vehicle for pathogens such as enteric viruses; nevertheless, the enteric viral circulation has been poorly studied in our region. This work aims to study the presence of five enteric virus, in parallel with operative parameters, in affluents and effluents of five wastewater treatment plants (WWTP) located in the Greater Metropolitan Area of Costa Rica in 2013. The viral pathogens studied were enterovirus, hepatitis A virus, rotavirus A, and norovirus GI and GII (determined by end point PCR technique); since basic control parameters analyzed were biochemical oxygen demand, chemical oxygen demand, suspended solids, settable solids, fats and oils, blue methylene active substances, and fecal coliforms. All wastewater treatment plants showed acceptable removal of physicochemical parameters, but with microbial parameters, its efficiency was low; all enteric viruses were detected year-round at effluents and influents of the WWTP, with a predominance during the dry season. The most frequently detected virus was rotavirus, followed by norovirus G1. In conclusion, the studied wastewater treatment plants are efficient to remove physicochemical parameters; the year circulation of enteric virus in Costa Rica was demonstrated, and finally, a wastewater treatment plant improvement it is necessary to achieved treated waters safety discharges.UCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias de la Salud::Instituto de Investigaciones en Salud (INISA

MIC of amoxicillin/clavulanate according to CLSI and EUCAST: Discrepancies and clinical impact in patients with bloodstream infections due to Enterobacteriaceae

Objectives: To compare results of amoxicillin/clavulanate susceptibility testing using CLSI and EUCAST methodologies and to evaluate their impact on outcome in patients with bacteraemia caused by Enterobacteriaceae. Patients and methods: A prospective observational cohort study was conducted in 13 Spanish hospitals. Patients with bacteraemia due to Enterobacteriaceae who received empirical intravenous amoxicillin/clavulanate treatment for at least 48h were included. MICs were determined following CLSI and EUCAST recommendations. Outcome variables were: failure at the end of treatment with amoxicillin/clavulanate (FEAMC); failure at day 21; and 30 day mortality. Classification and regression tree (CART) analysis and logistic regression were performed. Results: Overall, 264 episodes were included; the urinary tract was the most common source (64.7%) and Escherichia coli themost frequent pathogen (76.5%). Fifty-two isolates (19.7%) showed resistance according to CLSI and 141 (53.4%) according to EUCAST. The kappa index for the concordance between the results of both committees was only 0.24. EUCAST-derived, but not CLSI-derived, MICs were associated with failure when considered as continuous variables. CART analysis suggested a 'resistance' breakpoint of > 8/4mg/L for CLSI-derived MICs; it predicted FEAMC in adjusted analysis (OR=1.96; 95% CI: 0.98-3.90). Isolates with EUCAST-derived MICs > 16/2 mg/L independently predicted FEAMC (OR=2.10; 95%CI: 1.05-4.21) and failure at day 21 (OR=3.01; 95%CI: 0.93-9.67).MICs.32/2mg/Lwere only predictive of failure among patientswith bacteraemia from urinary or biliary tract sources. Conclusions: CLSI and EUCAST methodologies showed low agreement for determining the MIC of amoxicillin/clavulanate. EUCAST-derived MICs seemed more predictive of failure than CLSI-derived ones. EUCAST-derived MICs > 16/2 mg/L were independently associated with therapeutic failure

Espacios y destinos turísticos en tiempos de globalización y crisis

2 volúmenesXII Coloquio de Geografía del Turismo, Ocio y Recreación de la Asociación de Geógrafos Españoles. Colmenarejo (Madrid), del 17 al 19 de junio de 2010.Este libro ha sido editado con la colaboración económica del Ministerio de Ciencia e Innovación (ref. CS02010-10416-E)