Somatic Coliphages as an Operational Tool to Assess Loss of Bathing Water Quality after Heavy Rain Events.

Rapid population growth and coastal development has led to increased fecal contamination of coastal surface waters worldwide, enhancing the potential risk of waterborne human pathogens in bathing areas. More frequent heavy rainfall events, attributed to global warming, have further exacerbated the problem by causing sometimes sewer overflows into recreational waters. As traditional bacterial indicators have limited accuracy for predicting health risks associated with waterborne viruses, the additional use of viral indicators such as coliphages is recommended. In this study, we compared the behavior of bacterial and viral indicators of water quality at 10 Barcelona beaches during three bathing seasons, in dry conditions, and after four rainstorms that caused specific pollution events due to rain runoff with combined sewer overflows (CSO). Levels of all target indicators increased after the rainstorms, but compared to Escherichia coli and intestinal enterococci, somatic coliphages exhibited a slower decline and higher environmental persistence following a rain event. Daily continuous sampling carried out during the days following a rainstorm allowed not only the determination of the decay kinetics of each target indicator but also the day when the water quality recovered the values established in the current European regulation in approximately 2 -3 days after each CSO. These observations indicate that the combined use of bacterial and viral indicators can enhance the surveillance of microbial quality of bathing waters. Moreover, coliphages can swiftly provide insights into transient fecal pollution linked to rainfall episodes, thanks to available analytical techniques that enable same-day recommendations. The management of urban wastewater and recreational water regulations should consistently employ microbial indicators to address rainwater runoff or sewer overflows resulting from heavy rainfall.

Triacrylamide-Based Adhesives Stabilize Bonds in Physiologic Conditions.

In this study, an acrylamide-based adhesive was combined with a thiourethane-based composite to improve bond stability and reduce polymerization stress, respectively, of simulated composite restorations. The stability testing was conducted under physiologic conditions, combining mechanical and bacterial challenges. Urethane dimethacrylate was combined with a newly synthesized triacrylamide (TMAAEA) or HEMA (2-hydroxyethyl-methacrylate; control) to produce a 2-step total-etch adhesive system. Methacrylate-based composites (70 wt% silanized filler) were formulated, containing thiourethane oligomers at 0 (control) or 20 wt%. Standardized preparations in human third molars were restored; then, epoxy replicas were obtained from the occlusal surfaces before and after 7-d storage in water or with Streptococcus mutans biofilm, which was tested after storage in an incubator (static) or the bioreactor (mechanical challenge). Images were obtained from the replicas (scanning electron microscopy) and cross sections of the samples (confocal laser scanning microscopy) and then analyzed to obtain measurements of gap, bacterial infiltration, and demineralization. Microtensile bond strength of specimens stored in water or biofilm was assessed in 1-mm2 stick specimens. Data were analyzed with analysis of variance and Tukey's test (α = 0.05). HEMA-based materials had greater initial gap measurements, indicating more efficient bonding for the acrylamide materials. When tested in water, the triacrylamide-based adhesive had smaller gaps in the incubator or bioreactor. In the presence of biofilm, there was less difference among materials, but the acrylamide/thiourethane combination led to statistically lower gap formation in the bioreactor. HEMA and TMAAEA-based adhesives produced statistically similar microtensile bond strengths after being stored in water for 7 d, but after the same period with biofilm-challenged specimens, the TMAAEA-based adhesives were the only ones to retain the initial bond strength values. The use of a stable multiacrylamide-based adhesive led to the preservation of the resin-dentin bonded interface after a physiologically relevant challenge. Future studies will include a multispecies biofilm model.

Monitorización farmacocinética de antagonistas del factor de necrosis tumoral: utilidad clínica en enfermedad inflamatoria intestinal / Pharmacokinetics monitoring of tumour necrosis factor inhibitors: utility in inflammatory bowel disease management

Introducción: El fracaso de la terapia con biológicos en la enfermedad inflamatoria intestinal (EII) es un problema común. La monitorización farmacocinética es una herramienta que podría permitir optimizar estos tratamientos mediante la individualización de la posología en los diferentes escenarios clínicos. Objetivo: Describir las determinaciones de concentraciones séricas de infliximab y adalimumab en pacientes con EII y evaluar el impacto de las recomendaciones farmacocinéticas en la toma de decisiones clínicas y la evolución de los pacientes.Métodos: Estudio trasversal observacional retrospectivo de dos años de duración (2017-2018) de las determinaciones de inhibidores del factor de necrosis tumoral (anti-TNF) realizadas en pacientes del Servicio de Medicina Digestiva de un hospital general. Resultados: Se realizaron 133 determinaciones correspondientes a 66 sujetos (38% tratados con adalimumab; 62% infliximab). El principal motivo de monitorización (>55%) fue la presencia de enfermedad activa. Más de la mitad de los pacientes tenían concentraciones fuera del intervalo terapéutico (el 51% de adalimumab y 37% de infliximab presentaban una exposición subóptima). La recomendación de mantener la pauta posológica previa se propuso en <40% de los informes farmacocinéticos, siendo el grado de aceptación de la recomendación superior al 80% en todos los casos. Los datos analíticos de inflamación se redujeron tras la monitorización sin mostrar significación estadística en todos los casos.Conclusión: La mayoría de pacientes con EII tratados con anti-TNF presentan concentraciones de fármaco inadecuadas. La monitorización ha demostrado ser una herramienta útil para guiar la toma de decisiones terapéuticas y podría contribuir en la mejora de los parámetros clínicos de estos pacientes. (AU)

Introduction: The failure of biologic therapy in inflammatory bowel disease (IBD) is a common problem. The pharmacokinetic monitoring is a useful tool that allows to optimize these treatments.Objective: To describe the pharmacokinetic determinations of plasma concentrations of infliximab and adalimumab in patients with IBD and evaluate its impact on clinical decision.Method: Retrospective, observational study of two years duration (2017-2018) of the tumor necrosis factor (anti-TNF) inhibitor determinations performed in patients of the Digestive Service of a general hospital.Results: A total of 133 determinations were obtained about 66 subjects (38% adalimumab, 62% infliximab). The main reason (>55%) of the request for monitoring was the presence of active disease. More than half of the patients had concentrations outside the therapeutic range (51% of patients with adalimumab and 37% of those with infliximab had plasma levels below the lower range considered suboptimal). The maintenance of the previous dosage was recommended only in <40% of the monitoring and the degree of acceptance of the interventions were more than 80% in all cases. The analytical data of inflammation were reduced after monitoring without showing statistical significance in all cases.Conclusion: Standard anti-TNF regimen was not adequate to obtain range concentrations in more than half of analyzed cases. Monitoring has proven to be very useful in supporting clinical decision and it may promote an improvement in the clinical parameters of patients with IBD. (AU)

Global adherence to pharmacotherapeutic treatment in patients with multiple sclerosis / Adherencia global al tratamiento farmacoterapéutico en pacientes con esclerosis múltiple

Objective: To measure the indirect adherence of patients with multiple sclerosis (MS) to their pharmacological treatments, comparing medication for the disease itself with the other concomitant treatments that have been chronically prescribed.Methods: A cross-sectional, descriptive (November 2018) study at the Pharmacy Department of a spanish secondary hospital in Valencian Community. Study population is all those patients diagnosed with MS in its different variants, who are being treated with hospital dispensing drugs for a minimum of six months at the time of the study. The variable evaluated was the percentage of adherence of patients to their medication, which has been measured as the ratio between the doses of prescribed drugs and those dispensed; considering adherent to all patients with a percentage equal to or higher than 80%.Results: The study included 86 patients from the Pharmacy External Outpatient Unit. The average exposure time to drugs was 198.7±27.9 days, where the adherence to treatment of multiple sclerosis was 98.1±6.6% and that of the rest of the chronic medication concomitant 92.8±19.5%.Conclusions: Adherence to pharmacological treatments in this population has been very high. Patients have shown greater adherence to drugs dispensed at Pharmacy External Outpatient Unit, possibly because they consider the rest of medication less important and/or to treat milder comorbidities or pathologies. (AU)

Objetivos: Evaluar la adherencia indirecta de los pacientes con esclerosis múltiple (EM) a sus tratamientos farmacológicos, comparando la medicación para esta enfermedad entre los diferentes fármacos, y con el resto de tratamientos concomitantes prescritos por el médico de manera crónica.Material y métodos: Estudio descriptivo transversal (noviembre 2018) en el Servicio de Farmacia de un hospital secundario español de la Comunidad Valenciana. La población de estudio son todos aquellos pacientes diagnosticados con EM en sus diferentes variantes, que están siendo tratados con medicamentos dispensados desde el hospital, durante un periodo mínimo de seis meses. La variable evaluada fue el porcentaje de adherencia de los pacientes a su medicación, que se midió como la relación entre las dosis de los medicamentos prescritos y aquellos dispensados; considerando adherente a todos los pacientes con un porcentaje igual o superior al 80%.Resultados: Se incluyó a 86 pacientes desde la Unidad de Pacientes Externos del Servicio de Farmacia. El tiempo promedio de exposición a los fármacos para la EM fue de 198,7±27,9 días, observando una adherencia al tratamiento de 98,1±6,6% y una adherencia al resto de la medicación crónica concomitante de 92,8±19,5%.Conclusiones: La adherencia a los tratamientos farmacológicos en esta población ha sido muy alta. Los pacientes han mostrado una mayor adherencia a los medicamentos dispensados en la Unidad de Pacientes Externos del Servicio de Farmacia, posiblemente porque consideran que el resto de la medicación es menos importante y/o para tratar comorbilidades o patologías más leves. (AU)

Prevalência e distribuição espacial da ocorrência de helmintos em primatas não humanos de vida livre no estado do Rio de Janeiro, Brasil / [Prevalence and spatial distribution of the occurrence of helminths in free-living nonhuman primates in the State of Rio de Janeiro, Brazil]

Seiscentos primatas neotropicais foram submetidos a exames post mortem para avaliação da prevalência parasitária de helmintos gastrointestinais. Foram examinados 556 calitriquídeos (Callithrix spp.), 23 bugios (Alouatta guariba), 19 macacos-pregos (Sapajus nigritus), um mico-leão-dourado (Leontopithecus rosalia) e um mico-leão-da-cara-dourada (Leontopithecus chrysomelas). Do total de 600 animais, foram encontrados espécimes parasitos pertencentes aos filos Acanthocephala, Nemathelmintes e Platyhelminthes (classes Trematoda e Cestoda) em 110 primatas. A prevalência de primatas positivos para, pelo menos, uma espécie de helminto foi de 18,3% (110/600), sendo destes 83,6% (92/110) calitriquídeos, 8,2% (9/110) bugios, 6,4% (7/110) macacos-pregos, 0,9% (1/110) mico-leão-dourado e 0,9% (1/110) mico-leão-da-cara dourada. Em 80,4% (74/92) dos calitriquídeos foram encontrados nematoides Primasubulura sp. e em 1,1% (1/92) nematoides Trypanoxyuris callithrix, em 26,1% (24/92) acantocéfalos (Pachysentis sp.) e em 5,4% (5/92) digenéticos (Platynosomum sp.); em 77,8% (7/9) dos bugios foram encontrados nematoides (Trypanoxyuris minutus), em 11,1% (1/9) acantocéfalos (Pachysentis sp.) e em 11,1% (1/9) cestoides (Bertiella sp.); em 14,3% (1/7) dos macacos-pregos foram encontrados nematoides (Physaloptera sp.), em 28,6% (2/7) acantocéfalos (Prostenorchis sp.) e em 14,3% (1/7) digenéticos (Platynosomum sp.) e no mico-leão-da-cara-dourada foram encontrados acantocéfalos (Prostenorchis sp.). Foi realizado o georreferenciamento dos pontos de encontro dos cadáveres para pontuar a distribuição dos helmintos por região.(AU)

Six hundred neotropical primates underwent postmortem examinations to evaluate the parasitic prevalence of gastrointestinal helminths. Fifty-five callitrichids tamarins (Callithrix spp.), 23 howlers (Alouatta guariba), 19 nail monkeys (Sapajus nigritus), a golden lion tamarin (Leontopithecus rosalia) and a golden-faced lion tamarin (Leontopithecus chrysomelas) were examined. Parasitic specimens belonging to phylums Acantocephala, Nemathelmintes and Platyhelmintes (Trematoda and Cestoda Classes) were found. The prevalence of primates positive for at least one species of helminth was 18.3% (110/600), of which 83.6% (92/110) callitrichids, 8.2% (9/110) howler monkeys, 6.4% (7/110) capuchin monkeys, 0.9% (1/110) golden lion tamarin and 0.9% (1/110) golden faced lion tamarin. In 80.4% (74/92) of callitrichids nematodes (Primasubulura sp.) were found, and in 1.1% (1/92) nematodes (Trypanoxyuris callithricis), in 26.1% (24/92) acanthocephalus (Pachysentis sp.) and 5.4% (5/92) digenetics (Platynosomum sp.); in 77.8% (7/9) of howler monkeys presented nematodes (Trypanoxyuris minutus), 11.1% (1/9) acanthocephalus (Pachysentis sp.) and 11.1% (1/9) cestoids (Bertiella sp.); in 14.3% (1/7) of capuchin monkeys presented nematodes (Physaloptera sp.), 28.6% (2/7) acanthocephalus (Prostenorchis sp.) and 14.3% (1/7) digenetics (Platynosomum sp.) and in the golden-faced lion tamarin acanthocephalus (Prostenorchis sp.) were found. Georreferencing of the meeting points of the cadavers was performed in order to punctuate the distribution of helminths by region.(AU)

Modelling the seasonal impacts of a wastewater treatment plant on water quality in a Mediterranean stream using microbial indicators.

Faecal pollution modelling is a valuable tool to evaluate and improve water management strategies, especially in a context of water scarcity. The reduction dynamics of five faecal indicator organisms (E. coli, spores of sulphite-reducing clostridia, somatic coliphages, GA17 bacteriophages and a human-specific Bifidobacterium molecular marker) were assessed in an intermittent Mediterranean stream affected by a wastewater treatment plant (WWTP). Using Bayesian inverse modelling, the decay rates of each indicator were correlated with two environmental drivers (temperature and streamflow downstream of the WWTP) and the generated model was used to evaluate the self-depuration distance (SDD) of the stream. A consistent increase of 1-2 log10 in the concentration of all indicators was detected after the discharge of the WWTP effluent. The decay rates showed seasonal variation, reaching a maximum in the dry season, when SDDs were also shorter and the stream had a higher capacity to self-depurate. High seasonality was observed for all faecal indicators except for the spores of sulphite-reducing clostridia. The maximum SDD ranged from 3 km for the spores of sulphite-reducing clostridia during the dry season and 15 km for the human-specific Bifidobacterium molecular marker during the wet season. The SDD provides a single standardized metric that integrates and compares different contamination indicators. It could be extended to other Mediterranean drainage basins and has the potential to integrate changes in land use and catchment water balance, a feature that will be especially useful in the transient climate conditions expected in the coming years.

Dynamics of crAssphage as a human source tracking marker in potentially faecally polluted environments.

Recent studies have shown that crAssphage is abundant in human faecal samples worldwide. It has thus been postulated as a potential microbial source tracking (MST) marker to detect human faecal pollution in water. However, an effective implementation of crAssphage in water management strategies will depend on an understanding of its environmental dynamics. In this work, the abundance and temporal distribution of crAssphage was analysed in the effluent of wastewater treatment plants using different sewage treatments, and in two rivers (water and sediments) that differ in pollution impact and flow regime. Additionally, the influence of environmental conditions (temperature and rainfall) on the removal of the marker was studied along a river section, and natural inactivation was assessed by a mesocosms approach. Molecular and culture-based tools were used to compare crAssphage abundance and dynamics with those of bacteria and bacteriophages currently applied as global indicators (E. coli, somatic coliphages, Bacteroides GA17 bacteriophages, and the human-associated MST markers HF183 and HMBif). CrAssphage concentrations in sewage effluent and river samples were similar to those of HF183 and HMBif and higher than other general and/or culture-based indicators (by 2-3 orders of magnitude). Measurement of crAssphage abundance revealed no temporal variability in the effluent, although rainfall events affected the dynamics, possibly through the mobilisation of sediments, where the marker was detected in high concentrations, and an increase in diffuse and point pollution. Another factor affecting crAssphage inactivation was temperature. Its persistence was longer compared with other bacterial markers analysed by qPCR but lower than culturable markers. The results of this study support the use of crAssphage as a human source tracking marker of faecal pollution in water, since it has similar abundances to other molecular human MST markers, yet with a longer persistence in the environment. Nevertheless, its use in combination with infectious bacteriophages is probably advisable.

Pathogens, faecal indicators and human-specific microbial source-tracking markers in sewage.

The objective of this review is to assess the current state of knowledge of pathogens, general faecal indicators and human-specific microbial source tracking markers in sewage. Most of the microbes present in sewage are from the microbiota of the human gut, including pathogens. Bacteria and viruses are the most abundant groups of microbes in the human gut microbiota. Most reports on this topic show that raw sewage microbiological profiles reflect the human gut microbiota. Human and animal faeces share many commensal microbes as well as pathogens. Faecal-orally transmitted pathogens constitute a serious public health problem that can be minimized through sanitation. Assessing both the sanitation processes and the contribution of sewage to the faecal contamination of water bodies requires knowledge of the content of pathogens in sewage, microbes indicating general faecal contamination and microbes that are only present in human faecal remains, which are known as the human-specific microbial source-tracking (MST) markers. Detection of pathogens would be the ideal option for managing sanitation and determining the microbiological quality of waters contaminated by sewage; but at present, this is neither practical nor feasible in routine testing. Traditionally, faecal indicator bacteria have been used as surrogate indicators of general faecal residues. However, in many water management circumstances, it becomes necessary to detect both the origin of faecal contamination, for which MST is paramount, and live micro-organisms, for which molecular methods are not suitable. The presence and concentrations of pathogens, general faecal indicators and human-specific MST markers most frequently reported in different areas of the world are summarized in this review.

Faecal contamination in public pools in Barcelona province: Cryptosporidium spp. and bacterial indicators.

A study was conducted of the occurrence of Cryptosporidium in indoor heated public swimming pools and of three bacterial indicators (Escherichia coli, faecal enterococci and Clostridium perfringens) on pool surrounds. Although all examined pools adhered strictly to the Spanish regulations, the influence of several parameters related to water conditions, pool structure, users and location on the presence of protozoa and bacteria was analysed. Cryptosporidium was detected in 18.8% of pools in 60% of the five towns studied. The maximum concentration was 13 oocysts/L in one swimming pool and one Jacuzzi. The bacterial indicators' prevalence on pool surrounds was higher than 50%, being present in all of the towns. Plastic surfaces presented the lowest bacterial prevalence, whereas painted surfaces were 100% positive. No differences were observed for pool surrounds with autonomous or disabled users. Risk of cryptosporidiosis in pool vessels indicated that concentrations over 1 oocyst/10 L enhance the risk of infection, even in one exposure. Guidelines for managing faecal accidents and public information on the importance of good hygiene behaviours in and around swimming pools are recommended to limit oocysts' presence.

Bluephage: A rapid method for the detection of somatic coliphages used as indicators of fecal pollution in water.

The use of somatic coliphages as indicators of fecal and viral pollution in water and food has great potential due to the reliability, reproducibility, speed and cost effectiveness of methods for their detection. Indeed, several countries already use this approach in their water management policies. Although standardized protocols for somatic coliphage detection are available, user-friendly commercial kits would facilitate their routine implementation in laboratories. The new method presented here allows detection of up to 1 somatic coliphage in under 3.5 h, well within one working day. The method is based on a modified Escherichia coli strain with knocked-out uidB and uidC genes, which encode the transport of glucuronic acid inside cells, and overexpressing uidA, which encodes the enzyme ß-glucuronidase. The enzyme accumulated in the bacterial cells only has contact with its substrate after cell lysis, such as that caused by phages, since the strain cannot internalize the substrate. When the enzyme is released into the medium, which contains a chromogen analogous to glucuronic acid, it produces a change of color from yellow to dark blue. This microbiological method for the determination of fecal pollution via the detection of culturable microorganisms can be applied to diverse sample types and volumes for qualitative (presence/absence) and quantitative analysis and is the fastest reported to date.

A rapid validated UV-HPLC method for the simultaneous determination of the antiretroviral compounds darunavir and raltegravir in their dosage form.

OBJECTIVE: A rapid, simple and sensitive high-performance liquid chromatography (HPLC) method with ultraviolet detection has been developed for quantification of darunavir and raltegravir in their pharmaceutical dosage form. METHODS: The assay enables the measurement of both drugs with a linear calibration curve (R2= 0.999) over the concentration range 5-100 mg/L. The determination was performed on an analytical Tracer Excel 120 ODSB (15x0.4.6 cm) column at 35ºC. The selected wavelength was 254 nm. The mobile phase was a mixture of 0.037 M sodium dihydrogen phosphate buffer, acetonitrile and methanol (40:50:10, v/v/v) at a flow rate of 2.0 mL/min Nevirapine (50 mg/L) was used as internal standard. RESULTS: Accuracy, intra-day repeatability (n = 5), and inter-day precision (n = 3) were found to be satisfactory, being the accuracy from -4.33 to 3.88% and precisions were intra-day and inter-day, 0.25% and 4.42% respectively in case of darunavir. Raltegravir intra-day and inter-day precisions lower of 1.01 and 2.36%, respectively and accuracy values bet from -4.02 to 1.06%. CONCLUSIONS: Determination of the darunavir and raltegravir in their dosage form was done with a maximum deviation of 4%. This analytical method is rapid, easily implantable and offers good results.

Comparison of Two Concentration Methods for the Molecular Detection of Enteroviruses in Raw and Treated Sewage.

Human enteric viruses are a major causative agent of emerging waterborne diseases and constitute a serious public health concern. Environmental contamination occurs through discharge of waste materials from infected persons. Methods for viral detection should be developed to detect low infective dose of enteric viruses in environment. In this study, we aimed at comparing two concentration methods for the detection of naturally occurring enteroviruses in raw and treated sewage. In the first method, polyethylene glycol is used to concentrate viral particles from the collected samples. The second method is based on ultracentrifugation of viral particles at high speed (110,000×g). Genomes of enteroviruses were quantified by the quantitative real-time PCR method in raw and treated sewage samples. PEG-based method yielded higher genomic copies of enteric viruses (with an average of 5.9 log10 genomic copies/100 mL) when applied to raw sewage samples. While the ultracentrifugation assay in the second method decreases genomic copies number (with an average of 5.4 log10 genomic copies/100 mL). The recovery differences between the two methods were not significant when applied to clean samples (treated sewage). This could be explained by the presence of inhibitors, which interfere with qRT-PCR, in less quantity comparatively to raw sewage. PEG-based method would be more accurate for samples with high-organic matter load. This report emphasizes the importance of matrices nature on the recovery of enteroviruses from sewage samples. This should be taken into consideration for establishing standardized virological assays to ensure the virological quality control of discharged water in environment.

Direct ultrafiltration performance and membrane integrity monitoring by microbiological analysis.

The feasibility of substituting a conventional pre-treatment, consisting of dioxi-chlorination, coagulation/flocculation, settling and sand filtration, of a drinking water treatment plant (DWTP) by direct ultrafiltration (UF) has been assessed from a microbiological standpoint. Bacterial indicators, viral indicators and human viruses have been monitored in raw river, ultrafiltered and conventionally pre-treated water samples during two years. Direct UF has proven to remove bacterial indicators quite efficiently and to a greater extent than the conventional process does. Nevertheless, the removal of small viruses such as some small bacteriophages and human viruses (e.g. enteroviruses and noroviruses) is lower than the current conventional pre-treatment. Membrane integrity has been assessed during two years by means of tailored tests based on bacteriophages with different properties (MS-2, GA and PDR-1) and bacterial spores (Bacillus spores). Membrane integrity has not been compromised despite the challenging conditions faced by directly treating raw river water. Bacteriophage PDR-1 appears as a suitable microbe to test membrane integrity, as its size is slightly larger than the considered membrane pore size. However, its implementation at full scale plant is still challenging due to difficulties in obtaining enough phages for its seeding.

EU policy on sewage sludge utilization and perspectives on new approaches of sludge management.

This paper presents the current sewage sludge legislation in Europe and expected developments regarding the coming directives on the application of the "End-of-waste" criteria and on fertilizers. Discussion on sludge production and processing is also included. The Directive 86/278 has regulated the use in agriculture of residual sludge from domestic and urban wastewater. After 1986, this directive was transposed in the different member state legislation and currently the national limit values on heavy metals, some organic micropollutants and pathogens are placed in a rather wide range. This seems the inevitable consequence of different attitudes towards sludge management practices in the member states. The discussion by the European Joint Research Center (JRC) in Seville regarding application of end-of-waste criteria for compost and digestate has produced a final document (IPTS 2014) where sludge was excluded from the organic wastes admitted for producing an end-of-waste compost. Sludge processing in Europe seems addressed to different goals: sludge minimization, full stabilization and hygienization by thermal hydrolysis processes before anaerobic digestion, and on-site incineration by fluidized bed furnace. Thermophilic anaerobic digestion was applied with success on the Prague WWTP with a preliminary lysimeter centrifugation. Coming techniques, like wet oxidation and pyrolysis, are applied only on very few plants.

Effect of hygienization treatment on the recovery and/or regrowth of microbial indicators in sewage sludge.

AIMS: Escherichia coli (EC) is the primary indicator micro-organism in regulations for sewage sludge reuse. The aim of this work was to assess the ability of EC to enter and recover from a viable-but-not-culturable state (VBNC) after sludge hygienization treatments. METHODS AND RESULTS: The persistence of EC, somatic coliphages (SOMCPH), spores of sulphite-reducing clostridia (SRC) and Salmonella spp. was assessed in digested sludge after different pasteurization treatments and storage conditions. Pasteurization at 55°C produced EC-injured cells that were resuscitated during the first 24 h. Different sludge treatments altered the inactivation kinetics of EC, while SOMCPH and SRC did not resuscitate and showed lower die-off than EC. No regrowth was observed in stored sludge for up to 60 days. CONCLUSIONS: EC monitoring by culturable methods is not by itself a suitable method for assessing the hygienization achieved in sludge as EC can enter into VBNC from which it can recover during the first hours of storage. SIGNIFICANCE AND IMPACT OF THE STUDY: The regulations should indicate the time when monitoring of EC should be performed to avoid the period when EC can resuscitate from VBNC or add alternative microbial indicators, such as SOMCPH, which do not have a VBNC state.

Hygienization performances of innovative sludge treatment solutions to assure safe land spreading.

The present research aims at the evaluation of the hygienization performances of innovative sludge treatment processes applied for the separated treatment of secondary sludge. Namely, two digestion pretreatments (sonication and thermal hydrolysis) and two sequential biological processes (mesophilic/thermophilic and anaerobic/aerobic digestion) were compared to the mesophilic (MAD) and thermophilic anaerobic digestion (TAD). Microbial indicators (Escherichia coli, somatic coliphages and Clostridium perfringens spores) and pathogens (Salmonella and enteroviruses), which show different resistances to treatment processes, were monitored in untreated and treated sludge. Overall, microbial load in secondary sludge was shown to be similar or lower than previously reported in literature for mixed sludge. Notably, the anaerobic/aerobic digestion process increased the removal of E. coli and somatic coliphages compared to the simple MAD and always achieved the hygienization requirement (2-log-unit removal of E. coli) proposed by EU Commission in the 3rd Working Document on sludge (April 2000) for the use of treated sludges in agriculture with restriction on their application. The microbial quality limits for the unrestricted use of sludge in agriculture (no Salmonella in 50 g wet weight (WW) and E. coli <500 CFU/g) were always met when thermal digestion or pretreatment was applied; however, the required removal level (6-log-unit removal of E. coli) could not be assessed due to the low level of this microorganism in raw sludge. Observed levels of indicator removal showed a higher resistance of viral particles to thermal treatment compared with bacterial cells and confirmed the suitability of somatic coliphages as indicators in thermal treatment processes.

[Pharmaceutical intervention in duration of antimicrobial treatment at hospital ambit]. / Actuación farmacéutica en el control de la duración del tratamiento con antimicrobianos en el ámbito hospitalario.

OBJECTIVE: To estimate the acceptance of the pharmaceutical intervention in controlling duration of antimicrobial therapy and to evaluate their impact on optimizing the treatment. METHODS: Prospective observational study for two years in a General University Hospital. For the patients record, we followed non critical adult patients with antibiotic treatment. When the duration of antimicrobial treatment not complied with established criteria for each antibiotic and pathology, there was a communication with the physician, at which is recommended to assess the need for continue treatment. The acceptance of pharmaceutical intervention was collected and afterwards we analyzed the impact of this work by antimicrobial consumption and incidence of Clostridium difficile. RESULTS: . In 122 patients the pharmacist made a pharmaceutical intervention due to prolonged antibiotic treatment. The most prevalent antibiotics were ß-lactams, specifically meropenem. The intravenous administration was more frequent. In 77 cases it was decided to recommend the suspension of treatment, we conducted an orally prospective intervention at 70.15 % and the rest of interventions were written. Acceptance was 65.95 % and 65.00%, respectively. During the study period, the DDD of the antimicrobials decreased by 8.89% and expenditure on antimicrobials one 40.12%. The incidence of C. difficile was stable. CONCLUSIONS: . In a hospital, a pharmaceutical counselling program on the duration of antimicrobial therapy is well accepted by the prescriber physician, but it must be improved. The route of information does not affect the degree of acceptance. These actions could involve a reduction of antimicrobial consumption.

Effect of ultrasound, low-temperature thermal and alkali pre-treatments on waste activated sludge rheology, hygienization and methane potential.

Waste activated sludge is slower to biodegrade under anaerobic conditions than is primary sludge due to the glycan strands present in microbial cell walls. The use of pre-treatments may help to disrupt cell membranes and improve waste activated sludge biodegradability. In the present study, the effect of ultrasound, low-temperature thermal and alkali pre-treatments on the rheology, hygienization and biodegradability of waste activated sludge was evaluated. The optimum condition of each pre-treatment was selected based on rheological criteria (reduction of steady state viscosity) and hygienization levels (reduction of Escherichia coli, somatic coliphages and spores of sulfite-reducing clostridia). The three pre-treatments were able to reduce the viscosity of the sludge, and this reduction was greater with increasing treatment intensity. However, only the alkali and thermal conditioning allowed the hygienization of the sludge, whereas the ultrasonication did not exhibit any notorious effect on microbial indicators populations. The selected optimum conditions were as follows: 27,000 kJ/kg TS for the ultrasound, 80 °C during 15 min for the thermal and 157 g NaOH/kg TS for the alkali. Afterward, the specific methane production was evaluated through biomethane potential tests at the specified optimum conditions. The alkali pre-treatment exhibited the greatest methane production increase (34%) followed by the ultrasonication (13%), whereas the thermal pre-treatment presented a methane potential similar to the untreated sludge. Finally, an assessment of the different treatment scenarios was conducted considering the results together with an energy balance, which revealed that the ultrasound and alkali treatments entailed higher costs.

Quantification of relative proportions of intact cells in microbiological samples using the example of Cryptosporidium parvum oocysts.

UNLABELLED: The fast analysis of relative proportions of live and dead cells can be of great value whether for comparing inactivation efficiencies of different biocidal treatments or for monitoring organisms of interest in environmental samples. We introduce here a straightforward method to determine the percentage of intact cells based on treatment of samples with the viability dye propidium monoazide (PMA). PMA selectively enters membrane-damaged cells and suppresses their PCR detection through modification of their DNA. The study was performed using Cryptosporidium parvum oocysts as a model although the principle should be applicable to other organisms. Validation was performed with defined mixtures of live and heat-killed oocysts and by exposing oocysts to a heat stress gradient. The method correctly indicated increasingly lower proportions of intact cells with increasing temperatures. When comparing the loss of membrane integrity of UV-killed (40 mJ cm(-2) ) oocysts during storage in nonsterile tap water, results suggested that integrity declines slowly (over weeks) and at a rate comparable to non-UV-exposed oocysts. For all experiments, the amplification of longer DNA sequences was found beneficial. In the UV experiment, longer amplicons revealed not only higher sensitivity in excluding membrane-damaged oocysts, but also in excluding DNA with UV-induced damage. SIGNIFICANCE AND IMPACT OF THE STUDY: Whether in the context of microbial ecology or in an industrial context, many questions in microbiology are linked to microbial viability. As cultivation of micro-organisms can be long or may not be possible, fast methods to assess the numbers of live cells are in great demand. We present here a straightforward strategy to determine the relative proportions of intact cells. The PCR-based rapid method is expected to be useful where relative information is sufficient (e.g. for comparing the effect of different antimicrobial treatments on known numbers of micro-organisms) or when the presence of PCR inhibitors does not allow absolute quantification.

Alternative indicators for monitoring Cryptosporidium oocysts in reclaimed water.

With the widespread use of reclaimed water all over the world, there is a clear need to optimise its management in order to guarantee water safety. Model microorganisms (with either indicator or index function) are commonly used to assess risks related to the presence of enteric pathogens in water. Samples from five water reclamation plants located in Northeastern Spain were analysed to validate the use of three model microorganisms (Escherichia coli, somatic coliphages and spores of sulphite-reducing clostridia) as surrogates of Cryptosporidium total or infectious oocysts (TOO and IOO, respectively) in reclaimed water. Probability plots, simple and multiple linear regression and discriminant analyses were performed to assess their relationships. Results show that the detection of E. coli alone is not useful to model either the behaviour or concentrations of Cryptosporidium. However, discriminant analyses showed a high rate of correctly classified samples (91.9%) when E. coli and somatic coliphages data were used together to predict the presence/absence of IOO. Spores of sulphite-reducing clostridia (SRC) showed parallel reduction patterns and high correlation values (r = 0.76) with reductions in TOO. Furthermore, simple regression analyses of SRC and TOO in reclaimed water showed high correlation values (r = 0.85). Therefore, at the treatment plants studied, SRC can be considered to have good indicator and index functions for TOO. From the point of view of health protection, the use of SRC together with E. coli (which is mandatory in the current Spanish regulations) would satisfy the need for improved reclaimed water management.