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.

Analgesic Activity of 5-Acetamido-2-Hydroxy Benzoic Acid Derivatives and an In-Vivo and In-Silico Analysis of Their Target Interactions.

The design, synthesis, and evaluation of novel non-steroidal anti-inflammatory drugs (NSAIDs) with better activity and lower side effects are big challenges today. In this work, two 5-acetamido-2-hydroxy benzoic acid derivatives were proposed, increasing the alkyl position (methyl) in an acetamide moiety, and synthesized, and their structural elucidation was performed using 1H NMR and 13C NMR. The changes in methyl in larger groups such as phenyl and benzyl aim to increase their selectivity over cyclooxygenase 2 (COX-2). These 5-acetamido-2-hydroxy benzoic acid derivatives were prepared using classic methods of acylation reactions with anhydride or acyl chloride. Pharmacokinetics and toxicological properties were predicted using computational tools, and their binding affinity (kcal/mol) with COX-2 receptors (Mus musculus and Homo sapiens) was analyzed using docking studies (PDB ID 4PH9, 5KIR, 1PXX and 5F1A). An in-silico study showed that 5-acetamido-2-hydroxy benzoic acid derivates have a better bioavailability and binding affinity with the COX-2 receptor, and in-vivo anti-nociceptive activity was investigated by means of a writhing test induced by acetic acid and a hot plate. PS3, at doses of 20 and 50 mg/kg, reduced painful activity by 74% and 75%, respectively, when compared to the control group (20 mg/kg). Regarding the anti-nociceptive activity, the benzyl showed reductions in painful activity when compared to acetaminophen and 5-acetamido-2-hydroxy benzoic acid. However, the proposed derivatives are potentially more active than 5-acetamido-2-hydroxy benzoic acid and they support the design of novel and safer derivative candidates. Consequently, more studies need to be conducted to evaluate the different pharmacological actions, the toxicity of possible metabolites that can be generated, and their potential use in inflammation and pain therapy.

Bacteriophages in sewage: abundance, roles, and applications.

The raw sewage that flows through sewage systems contains a complex microbial community whose main source is the human gut microbiome, with bacteriophages being as abundant as bacteria or even more so. Phages that infect common strains of the human gut bacteriome and transient bacterial pathogens have been isolated in raw sewage, as have other phages corresponding to non-sewage inputs. Although human gut phages do not seem to replicate during their transit through the sewers, they predominate at the entrance of wastewater treatment plants, inside which the dominant populations of bacteria and phages undergo a swift change. The sheer abundance of phages in the sewage virome prompts several questions, some of which are addressed in this review. There is growing concern about their potential role in the horizontal transfer of genes, including those related with bacterial pathogenicity and antibiotic resistance. On the other hand, some phages that infect human gut bacteria are being used as indicators of fecal/viral water pollution and as source tracking markers and have been introduced in water quality legislation. Other potential applications of enteric phages to control bacterial pathogens in sewage or undesirable bacteria that impede the efficacy of wastewater treatments, including biofilm formation on membranes, are still being researched.

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.

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.

Use of mini implants to replace a missing tooth in a growing patient: a six-year follow up case report.

BACKGROUND: Loss of anterior permanent teeth can have a detrimental effect on children's quality of life and the therapeutic approaches are always a challenge. From a physiologic and psychological standpoint, the use of dental implants has several advantages in replacing missing teeth. However, several studies have shown that conventional implants are not indicated in patients that are still growing, since they interfere with the sagittal and transversal growth of the maxilla. Recent literature has suggested that Mini Dental Implants (MDIs) can be successfully applied in growing patients, without interfering with the normal craniofacial growth process. CASE REPORT: This report describes the replacement of missing teeth in a 10-year-old patient by a mini implant of 1.8 mm diameter. After a six-year follow-up period, the mini implant did not follow the regular growth process of the maxilla, resulting in functional and aesthetic complications. Removal of the mini implant entailed some difficulties.

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.

Bacteriophages as indicators of human and animal faecal contamination in raw and treated wastewaters from Tunisia.

AIMS: We aimed at quantifying bacteriophages in raw and treated wastewaters of human and animal origin in Tunisia to assess their usefulness for tracking the origin of faecal pollution and in the follow-up of effectiveness of water treatments process. METHODS AND RESULTS: The concentrations of bacteriophages in wastewater samples were determined by double layer agar technique. Somatic coliphages and F-specific RNA bacteriophages were present in all types of samples in high concentrations. The values of Escherichia coli were variable depending on geographical location. On the other hand, bacteriophages infecting strain GA17 were detected preferably when human faecal contamination was occurred. CONCLUSIONS: Bacteriophages appear as a feasible and widely applicable manner to detect faecal contamination in Tunisia. On the other hand, phages infecting GA17 could be good markers for tracking the origin of faecal pollution in the area studied. SIGNIFICANCE AND IMPACT OF THE STUDY: The reuse of treated wastewaters can be a solution to meet the needs of water in the geographical area of study. Bacteriophages seem to predict differently the presence of faecal contamination in water than bacterial indicators. Consequently, they can be a valuable additional tool to improve water resources management for minimizing health risks.

Insertion Torque of Mini Dental Implants and their Failure Risk after Immediate Loading / Torque de Inserción de Mini Implantes Dentales y su Riesgo de Fracaso cuando son Cargados Inmediatamente

A minimum insertion torque has been suggested for immediate function of regular diameter implants. Even when there is a growing tendency to use Mini Dental Implants (MDIs) for immediate function of dentures, there is no clinical data concerning MDIs insertion torque, nor its influence on implant failure. The aim of this prospective study was to assess the insertion torque of immediate loaded MDIs in edentulous patients and its association with risk failure. Ninety MDIs were placed in the anterior mandible of 45 edentulous patients, two per patient. The insertion torque was recorded with an electronic device and failures were documented during two years follow-up. All implants were immediately loaded with overdentures, using ball (44/90) or bar (46/90) attachments. A Kaplan­Meier survival probability estimator and a fitted multiple Cox regression model were performed to establish the influence of insertion torque and other clinical parameters on implant risk failure. The average insertion torque of 90 MDIs was 12.5±7.8 Ncm. A cumulative survival rate of 94.2% (5/90) was found by means of the Kaplan-Meier estimation. The Cox proportional-hazards regression model showed no association between insertion torque and MDIs failure. MDIs have much lower insertion torque than average conventional implants. The insertion torque of MDIs immediately loaded with mandibular dentures, seems not to be a risk factor associated to failure, at two years follow-up.

Un mínimo torque de inserción ha sido sugerido para realizar carga inmediata de implantes de diámetro regular. Aún cuando existe una tendencia creciente a utilizar Mini Implantes Dentales (MID) para la función inmediata de las prótesis totales, no hay datos clínicos relativos a los valores de torque de inserción de estos, ni tampoco de su influencia en el fracaso del implante. El objetivo de este estudio prospectivo fue evaluar el torque de inserción de mini implantes dentales con carga inmediata en pacientes desdentados y su asociación con el riesgo de fracaso. Noventa MIDs fueron colocados en la mandíbula anterior de 45 pacientes desdentados, dos por paciente. El torque de inserción fue obtenido con un dispositivo electrónico y los fracasos fueron documentadas a dos años de seguimiento. Todos los implantes fueron cargados inmediatamente con sobredentaduras, usando un sistema de retención de bola (44/90) o barra (46/90). Se utilizó el estimador de probabilidad de sobrevida de Kaplan-Meier y el modelo riesgos proporcionales de Cox ajustado para establecer la influencia de torque de inserción y otros parámetros clínicos sobre el riesgo de fracaso del implante. El promedio de torque de inserción de 90 MIDs fue 12,5±7,8 Ncm . Los MIDs mostraron a través de la estimación de Kaplan-Meier, una tasa de supervivencia acumulada de 94,2% (5/90). El modelo de riesgos proporcionales de Cox reveló que no existe asociación entre el valor de torque de inserción y el fracaso MIDs. MIDs tienen un valor de torque de inserción mucho más bajo que los implantes convencionales. El torque de inserción de MIDs cargados inmediatamente con prótesis mandibulares, no parece ser un factor de riesgo asociado al fracaso, a dos años de seguimiento.

Bases genéticas y moleculares del cáncer colorrectal / Genetic and molecular basis in colorectal cancer

El cáncer colorrectal (CCR) es la tercera neoplasia más frecuente a un nivel mundial. El riesgo de CCR depende de factores genéticos y ambientales que interactúan en el proceso de carcinogénesis mediante mecanismos epigenéticos. Aunque sólo el 25 % de los CCR presenta agregación familiar, en todos los casos de CCR entre los que se incluyen los esporádicos se producen mutaciones genéticas que desencadenan tres tipos distintos de vías moleculares de carcinogénesis que influyen en un comportamiento tumoral diferente. El conocimiento de los genes y las vías moleculares implicados en el desarrollo del CCR tiene unas aplicaciones clínicas debido a que ayuda a definir el pronóstico, a elaborar un consejo genético en caso de síndromehereditario y a diseñar un plan terapéutico más específico de acuerdo a las características moleculares del tumor. Por otro lado, según la teoría de las células madre, una célula madre del epitelio colónico con una acumulación de aberraciones genéticas puede ser la célula origen del CCR. El diseño de nuevos fármacos dirigidos contra marcadores de las células madre del CCR pueden potenciar la respuesta de la quimioterapia convencional y disminuir las recurrencias locales y metástasis a distancia (AU)

Colorectal cancer is the third most common cancer wordlwide. Relative CRC risk is defined by genetic and environmental factors contributing to carcinogenesis by epigenetic mechanisms. Though only a 25 % CRC has been determined to have a familiar agregation with an important genetic component, all cases of CRC including sporadic ones present genetic aberrations that initiate three distinct molecular pathways of carcinogenesis implicating in different tumoral behaviours. Investigations about genes and molecular pathways implicated in the development of CRC have clinic aplications due to it could contribute to determine prognosis, elaborate a genetic conseling in case of familiar agregation and design therapeutic strategies according with molecular characteristics of tumor. On the other hand, A stem cell located in colonic epithelium with an acumulation of genetic mutations could be the cell that initiates the origin of CRC. The design of stem cell-targeted drugs could enhance reponsiveness to traditional therapeutic strategies and reduce local recurrence and metástasis (AU)

Balancing hygienization and anaerobic digestion of raw sewage sludge.

The anaerobic digestion of raw sewage sludge was evaluated in terms of process efficiency and sludge hygienization. Four different scenarios were analyzed, i.e. mesophilic anaerobic digestion, thermophilic anaerobic digestion and mesophilic anaerobic digestion followed by a 60 °C or by an 80 °C hygienization treatment. Digester performance (organic matter removal, process stability and biogas yield) and the hygienization efficiency (reduction of Escherichia coli, somatic coliphages and F-specific RNA phages) were the main examined factors. Moreover, a preliminary economical feasibility study of each option was carried out throughout an energy balance (heat and electricity). The obtained results showed that both thermophilic anaerobic digestion and mesophilic anaerobic digestion followed by a hygienization step were able to produce an effluent sludge that fulfills the American and the European legislation for land application. However, higher removal efficiencies of indicators were obtained when a hygienization post-treatment was present. Regarding the energy balance, it should be noted that all scenarios have a significant energy surplus. Particularly, positive heat balances will be obtained for the thermophilic anaerobic digestion and for the mesophilic anaerobic digestion followed by 60 °C hygienization post-treatment if an additional fresh-sludge/digested sludge heat exchanger is installed for energy recovery.

Feasibility of methods based on nucleic acid amplification techniques to fulfil the requirements for microbiological analysis of water quality.

Molecular methods based on nucleic acid recognition and amplification are valuable tools to complement and support water management decisions. At present, these decisions are mostly supported by the principle of end-point monitoring for indicators and a small number of selected measured by traditional methods. Nucleic acid methods show enormous potential for identifying isolates from conventional culture methods, providing data on cultivable and noncultivable micro-organisms, informing on the presence of pathogens in waters, determining the causes of waterborne outbreaks, and, in some cases, detecting emerging pathogens. However, some features of water microbiology affect the performance of nucleic acid-based molecular techniques and thus challenge their suitability for routine water quality control. These features include the variable composition of target water samples, the generally low numbers of target micro-organisms, the variable water quality required for different uses and the physiological status or condition of such micro-organisms. The standardization of these molecular techniques is also an important challenge for its routine use in terms of accuracy (trueness and precision) and robustness (reproducibility and reliability during normal usage). Most of national and international water regulations recommend the application of standard methods, and any new technique must be validated respect to established methods and procedures. Moreover, molecular methods show a high cost-effectiveness value that limits its practicability on some microbial water analyses. However, new molecular techniques could contribute with new information or at least to supplement the limitation of traditional culture-based methods. Undoubtedly, challenges for these nucleic acid-based methods need to be identified and solved to improve their feasibility for routine microbial water monitoring.

Quantification of Shiga toxin 2-encoding bacteriophages, by real-time PCR and correlation with phage infectivity.

AIMS: To evaluate a qPCR-based protocol for the enumeration of Shiga toxin (Stx) 2 phages and to compare the results of qPCR with the number of infective Stx phage particles. METHODS AND RESULTS: An approach based on qPCR was applied to count Stx phages in five phage lysates of known titre. The number of viral particles from each phage lysate was determined by electron microscopy using latex spheres. The infectivity of the Stx phages was evaluated onto three bacterial host strains, by double agar layer assay and plaque blot hybridization. The number of phage particles detected by electron microscopy correlates with the number calculated by qPCR in all the phages assayed. The number of infectious phages was from 1 to 3 log10 units below the numbers obtained by qPCR and electron microscopy. CONCLUSIONS: The approach allows accurate quantification of Stx phages with a high recovery. The number of infectious phages is always below the number of phage particles detected by qPCR. SIGNIFICANCE AND IMPACT OF THE STUDY: The qPCR method is a good approach to enumerate Stx phages. However, these results should be carefully considered when related to the number of infectious phages for each lysate that could be applied in real samples, because values of infectious particles are always below the number of Stx phages detected by qPCR.

The effect of maximum bite force on marginal bone loss of mini-implants supporting a mandibular overdenture: a randomized controlled trial.

OBJECTIVES: To evaluate the effect of maximum bite force (mBF) on marginal bone loss (MBL) around mini-implants in edentulous patients wearing mandibular overdentures with two retention systems: ball and bar. MATERIAL AND METHODS: Forty-five totally edentulous patients were selected from a public health center. All of them received two mini-implants (1.8 x 15 mm; Sendax) in the anterior mandible using a minimally invasive technique. A single randomization was performed to allocate the patients in two groups. Group I (n=22) received two single ball-type mini-implants and Group II (n=23) received two mini-implants splinted with a prefabricated bar. The mBF was recorded using a press-sensitive sheet Dental Prescale (Fuji) and MBL using standardized radiographs of each mini-implant at the baseline and 5, 7, 10, and 15 months after surgery; the values were compared between groups. RESULTS: Two members of Group I failed to complete the study, decreasing the number of participants to 20. There was no relationship between the mBF and the MBL of the mini-implants (Spearman's rhor(s)=0.147; P=0.378). At the 15-month follow-up, the average mBF for Group I (ball) was 247.53 +/- 132.91 N and that of Group II (bar) only 203.23 +/- 76.85 N (Mann-Whitney test; P=0.586). The MBL values were also higher for Group I (1.40 +/- 1.02 mm) than Group II (0.84 +/- 0.66 mm) during the entire 15-month follow-up period (Mann-Whitney test; P=0.077). CONCLUSIONS: No relationship was found between mBF and MBL for patients wearing overdentures retained on mini-implants using bar or ball attachment systems.

Is the replication of somatic coliphages in water environments significant?

Somatic coliphages are amid several groups of bacteriophages that have been suggested as indicators in water quality assessment. One of the limitations frequently endorsed to somatic coliphages as indicators is that they can replicate in the water environment. This review intends to evaluate the significance of this potential replication. In view of: the threshold densities of somatic coliphages and host bacteria needed for productive infection to occur, the densities of both host cells supporting somatic coliphages replication and these phages in water environments, and the poor contribution of lysogenic induction to the free somatic coliphage numbers in water, it can be concluded that replication of somatic coliphages in waters is very unlikely. Consequently, the contribution of replication in the environment of somatic coliphages is expected to have a non-noticeable influence on the numbers of somatic coliphages detected in water environments. Thus, the replication in the environment should not be argued as a limitation to the use of somatic coliphages as indicators.

Pancreatitis aguda hidatídica / No disponible

No disponible

The CI repressors of Shiga toxin-converting prophages are involved in coinfection of Escherichia coli strains, which causes a down regulation in the production of Shiga toxin 2.

Shiga toxins (Stx) are the main virulence factors associated with a form of Escherichia coli known as Shiga toxin-producing E. coli (STEC). They are encoded in temperate lambdoid phages located on the chromosome of STEC. STEC strains can carry more than one prophage. Consequently, toxin and phage production might be influenced by the presence of more than one Stx prophage on the bacterial chromosome. To examine the effect of the number of prophages on Stx production, we produced E. coli K-12 strains carrying either one Stx2 prophage or two different Stx2 prophages. We used recombinant phages in which an antibiotic resistance gene (aph, cat, or tet) was incorporated in the middle of the Shiga toxin operon. Shiga toxin was quantified by immunoassay and by cytotoxicity assay on Vero cells (50% cytotoxic dose). When two prophages were inserted in the host chromosome, Shiga toxin production and the rate of lytic cycle activation fell. The cI repressor seems to be involved in incorporation of the second prophage. Incorporation and establishment of the lysogenic state of the two prophages, which lowers toxin production, could be regulated by the CI repressors of both prophages operating in trans. Although the sequences of the cI genes of the phages studied differed, the CI protein conformation was conserved. Results indicate that the presence of more than one prophage in the host chromosome could be regarded as a mechanism to allow genetic retention in the cell, by reducing the activation of lytic cycle and hence the pathogenicity of the strains.