New Perspectives in Monitoring Drinking Water Microbial Quality

The safety of drinking water is evaluated by the results obtained from faecal indicators during the stipulated controls fixed by the legislation. However, drinking-water related illness outbreaks are still occurring worldwide. The failures that lead to these outbreaks are relatively common and typically involve preceding heavy rain and inadequate disinfection processes. The role that classical faecal indicators have played in the protection of public health is reviewed and the turning points expected for the future explored. The legislation for protecting the quality of drinking water in Europe is under revision, and the planned modifications include an update of current indicators and methods as well as the introduction of Water Safety Plans (WSPs), in line with WHO recommendations. The principles of the WSP approach and the advances signified by the introduction of these preventive measures in the future improvement of dinking water quality are presented. The expected impact that climate change will have in the quality of drinking water is also critically evaluated

¿Se comunican los virus?

In 2017, several israeli virologists have demonstrated that bacterium-targeting virases, known as bacteriophages or phages, showed a collaborative aspect to a developmental decision-making process. In this case, intercellular communication occurs through the use of a viral peptide, named arbitrium and, in addition, two gene-products are involved, an intracellular-receptor and a negative regulator. The arbitrium system provides an elegant mechanism for a phage particle to estimate the amount of recent previous infections and hence decide whether to employ the lytic or lysogenic cycle. Intercellular molecular communication between viruses had not been observed before, and the study described here illuminates a previously unknown mechanism of viral action.En 2017 investigadores israelíes demostraron que algunos virus bacterianos, denominados bacteriófagos o simplemente fagos, mostraban una colaboración para desarrollar un proceso de «toma de decisión». En este caso, la comunicación intracelular se produce por un péptido vírico, denominado arbitrium, y además, están implicados dos péptidos más, un receptor intracelular y un regulador negativo. El sistema arbitrium representa un elegante mecanismo para que una partícula fágica estime la cantidad de progenie vírica de un previo proceso infectivo y así poder decidir si produce un ciclo lítico o lisogénico en la célula hospedadora. La comunicación intracelular entre virus no ha sido descrita previamente, y este estudio aclara mecanismos de replicación vírica desconocidos hasta ahor

Enumeration and isolation of viral particles from oligotrophic marine environments by tangential flow filtration

A method for concentrating, enumerating and isolating viral particles from marine water samples was developed and evaluated. The method consists of a concentration step by a tangential flow filtration (TFF) system, ultrafiltration by centrifugal concentrator, and visualization by transmission electron microscopy (TEM). This procedure allows to reduce volumes of ca. 2 l of seawater to 10–20 μl, which can be dispensed on electron microscopy grids to count total viral particles. This method allows the recovery of small numbers of viral particles from oligotrophic seawater samples, in which viral numbers ranged from 105 to 106 viral particles/ml. The tangential flow filtration system was evaluated as quantitative technique using suspensions of two different bacteriophages (T6 and ΦX174) in autoclaved seawater. Recovery rates varied depending on both the viral morphology and flow rate; recovery percentages reached 117.4% for T6 and 60.6% for ΦX174 using low flow rate

Detección y cuantificación del virus de la enfermedad de linfocistis mediante ensayo ICC-RT-PCR (integrated cell culture-RT-PCR)

La enfermedad de linfocistis es la enfermedad de etiología viral más frecuentemente detectada en la acuicultura marina europea, siendo la principal patología de origen vírico descrita en doradas cultivadas. El agente etiológico de esta enfermedad es el virus de la enfermedad de linfocistis (LCDV), miembro del género Lymphocystivirus, perteneciente a la familia Iridoviridae. Se han desarrollado diversos protocolos de PCR y PCR a tiempo real que permiten la detección y cuantificación del LCDV en diversas muestras, si bien no aportan ninguna indicación de la infectividad de los virus detectados. La detección de partículas víricas infectivas requiere la utilización de cultivos celulares, pero en el caso del LCDV la observación de efectos citopáticos (CPE) es difícil y a menudo está sujeta a subjetividad, especialmente en muestras con baja carga vírica. Por este motivo, en el presente trabajo se ha desarrollado un ensayo de ICC-RT-PCR (Integrated Cell Culture-RT-PCR) que permite la detección de partículas infectivas del LCDV. Este ensayo se ha aplicado en combinación con el método del número más probable (NMP) para la determinación del título infectivo en cultivos celulares. El protocolo de ICC-RT-PCR desarrollado permitió la detección de mRNA viral a partir de células SAF-1 inoculadas con un título infectivo de LCDV de 0,1 TCID50/ml, procesadas a los 5 d p.i, mientras que el límite de detección mediante observación de CPE fue de 10 TCID50/ml a 14 d p.i. La sensibilidad de la técnica he permitido la detección de partículas infectivas del LCDV en ejemplares de dorada asintomáticos, donde no se observaron CPE en cultivos celulares inoculados en paralelo y mantenidos hasta 14 d p.i. Este protocolo también se ha aplicado para la determinación del título infectivo de diferentes aislados víricos obtenidos a partir de peces enfermos, reduciéndose de forma considerable el tiempo necesario para realizar la titulación en comparación con el método de la dosis infectiva 50% en cultivos celulares (TCID50) (5 d versus 14-21 d, respectivamente). Así mismo, se han titulado stocks víricos obtenidos en cultivos celulares, donde la carga vírica es inferior al límite de detección del ensayo de observación de CPE. En conclusión, el protocolo de ICC-RT-PCR desarrollado es una técnica sensible, rápida y útil para la detección y cuantificación de LCDV infectivos, lo que la convierte en una herramienta adecuada para el estudio de esta patología viral.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tec

Are the defined substrate-based methods adequate to determine the microbiological quality of natural recreational waters?

Monitoring the microbiological quality of water used for recreational activities is very important to human public health. Although the sanitary quality of recreational marine waters could be evaluated by standard methods, they are time-consuming and need confirmation. For these reasons, faster and more sensitive methods, such as the defined substrate-based technology, have been developed. In the present work, we have compared the standard method of membrane filtration using Tergitol-TTC agar for total coliforms and Escherichia coli, and Slanetz and Bartley agar for enterococci, and the IDEXX defined substrate technology for these faecal pollution indicators to determine the microbiological quality of natural recreational waters. ISO 17994: 2004 standard was used to compare these methods. The IDEXX for total coliforms and E. coli, Colilert (R), showed higher values than those obtained by the standard method. Enterolert (R) test, for the enumeration of enterococci, showed lower values when compared with the standard method. It may be concluded that more studies to evaluate the precision and accuracy of the rapid tests are required in order to apply them for routine monitoring of marine and freshwater recreational bathing areas. The main advantages of these methods are that they are more specific, feasible and simpler than the standard methodology

Revisiting the genus Photobacterium: taxonomy, ecology and pathogenesis

The genus Photobacterium, one of the eight genera included in the family Vibrionaceae, contains 27 species with valid names and it has received attention because of the bioluminescence and pathogenesis mechanisms that some of its species exhibit. However, the taxonomy and phylogeny of this genus are not completely elucidated; for example, P. logei and P. fischeri are now considered members of the genus Aliivibrio, and previously were included in the genus Vibrio. In addition, P. damselae subsp. piscicida was formed as a new combination for former Vibrio damsela and Pasteurella piscicida. Moreover, P. damselae subsp. damselae is an earlier heterotypic synonym of P. histaminum. To avoid these incovenences draft and complete genomic sequences of members of Photobacterium are increasingly becoming available and their use is now routine for many research laboratories to address diverse goals: species delineation with overall genomic indexes, phylogenetic analyses, comparative genomics, and phenotypic inference. The habitats and isolation source of the Photobacterium species include seawater, sea sediments, saline lake waters, and a variety of marine organisms with which the photobacteria establish different relationships, from symbiosis to pathogenic interactions. Several species of this genus contain bioluminescent strains in symbiosis with marine fish and cephalopods; in addition, other species enhance its growth at pressures above 1 atmosphere, by means of several high-pressure adaptation mechanisms and for this, they may be considered as piezophilic (former barophilic) bacteria. Until now, only P. jeanii, P. rosenbergii, P. sanctipauli, and the two subspecies of P. damselae have been reported as responsible agents of several pathologies on animal hosts, such as corals, sponges, fish and homeothermic animals. In this review we have revised and updated the taxonomy, ecology and pathogenicity of several members of this genus. [Int Microbiol 20(1): 1-10 (2017)]Keywords: Photobacterium · taxonomy · symbiosis · pathogenesis · virulence factor

Determinación de órganos diana para la multiplicación y persistencia del virus de la enfermedad de linfocistis (LCDV) en dorada (Sparus aurata, L.)

La enfermedad de linfocistis es la única patología de etiología viral descrita en dorada cultivada En la cuenca mediterr nea, la prevalencia es cercana al 100 , ocasionando graves p rdidas económicas debido a la imposibilidad de comercializar los peces afectados En el presente trabajo se ha abordado el estudio de la patog nesis del virus de la enfermedad de linfocistis (LCDV) en dorada, además se han establecido los órganos implicados en la multiplicación vírica Para ello, se ha diseñado un protocolo de hibridación in situ empleando sondas RNA marcadas con digoxigenina dirigidas contra el gen que codifica la proteína principal de la cápside (MCP) viral, y se ha evaluado en poblaciones de dorada. En paralelo, se ha procedido a la cuantificación del número de copias de genoma viral por PCR a tiempo real y cuantificación relativa de la transcripción del gen que codifica la MCP viral mediante qRT-PCR. Los resultados obtenidos indican que el LCDV establece una infección sistémica en alevines de dorada, pudiendo detectarse señal de hibridación tanto en órganos internos (hígado, bazo, riñón) como en músculo y aleta. También se han observado diversos daños histopatológicos en animales enfermos, mientras que en animales recuperados de la enfermedad estos daños parecen revertir, aunque en estos animales la infección persiste, si bien sólo a niveles detectables mediante PCR a tiempo real.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Comparative Melodic Analysis of A Cappella Flamenco Cantes.

Background in ethnomusicology and music analysis. A cappella singing styles (called cantes in the flamenco jargon) are among the most fundamental song styles within the flamenco repertoire. Until very recently, flamenco singers have been only using oral transmission to learn them. Because of this form of diffusion, melody has become one of the main musical facets to be listened to, remembered, elaborated and spread in flamenco singing styles. Moreover, melody has helped flamenco enthusiasts to remember and identify variants of a particular style or genre. A frequent discussion and unanswered question among flamenco scholars is how to quantify the similarity between two melodies, and how to use this similarity measure to differentiate different styles and variants among performers, and to study the roots and evolution of flamenco styles

Potential risk of Artemia sp as a transmission vector of lymphocystis disease virus (LCDV)

4 páginas, 1 figura. XI Congreso Nacional de Acuicultura (Vigo, 24-28 septiembre 2007). Ed. Antonio Cerviño Eiroa, Alejandro Guerra Díaz y Carmen Pérez Acosta.[EN] Cysts and naupliis of Artemia sp. were analysed. Specific nested-PCR revealed lymphocystis disease virus (LCDV) genome. Infective viral particle has been observed by CPE development in inoculated cell cultures. Viral genome was found by in situ hybridization in nauplii and adults of natural infected artemia, as in nauplii of bath challenged artemia. No morphological damages have been observed. Artemia is a bio-accumulator of fish pathogens, with a possible role as environmental reservoir of fish pathogens. These results have shown the risk of artemia as a source of viral pathogens to fish larvae.[ES] En el presente trabajo se han realizado estudios virológicos en distintos lotes de quistes comerciales de artemias utilizados como alimento de larvas de peces marinos en piscifactorías. Mediante nested-PCR se ha detectado genoma del virus de linfocistis (LCDV) en homogeneizados de quistes y nauplios de artemia. La existencia de partículas víricas infectivas en estos homogeneizados se ha demostrado mediante la aparición de efectos citopáticos en cultivos celulares. Los ensayos de hibridación in situ han demostrado la existencia del LCDV en artemias infectadas naturalmente, así como en artemias inoculadas mediante baño, sin observarse alteraciones morfológicas. Las artemias actúan por tanto como bioacumuladores, pudiendo desempeñar un papel importante como reservorios ambientales de patógenos de peces. Estos resultados ponen de manifiesto el riesgo potencial de las artemias como fuente de patógenos víricos en estadios larvarios.Irene Cano es contratada postdoctoral en el ICMAN.CSIC dentro del Fondo social europeo- I3P-CSIC, en el marco del proyecto AGL2006-17777-C03-02/ACU (IP: Carmen Sarasquete).Peer reviewe

Co-occurrence of viral and bacterial pathogens in disease outbreaks affecting newly cultured sparid fish

Several microbial disease outbreaks in farm stocks of newly cultured sparid fish species, such as common seabream, redbanded seabream, and white seabream, were recorded from 2004 to 2006. This study describes the isolation and characterization of the potential causative agents, either bacteria or viruses, of these outbreaks. The isolated bacterial strains were characterized according to traditional taxonomical analyses and sequencing of a 16S rDNA fragment. Most bacteria were identified as Vibrio spp. and Photobacterium damselae subsp. damselae. The development of cytopathic effects (CPE) on different fish cell lines, the application of specific nested-PCR tests for infectious pancreatic necrosis virus (IPNV), viral nervous necrosis virus (VNNV) and viral hemorrhagic septicemia virus (VHSV), and subsequent sequence analyses were used for virus detection and identification. VNNV, related to the striped jack neural necrosis virus (SJNNV) genotype, and VHSV, related to the genotype Ia, were the only viruses detected. VNNV was isolated from the three fish species under study in five different outbreaks, whereas VHSV was isolated from common seabream and white seabream during two of these outbreaks. IPNV was not detected in any case. [Int Microbiol 2007; 10(3):193-199