Procedimiento de preparación, conservación, y uso en peces, del probiótico SHEWANELLA PUTREFACIENS PDP 11 para el control de enfermedades y la mejora en el crecimiento

Número de solicitud 201100385Procedimiento de preparación, conservación, y uso en peces, del probiótico Shewanella putresfaciens Pdp11 para el control de enfermedades y la mejora en el crecimiento. Preferentemente, el probiótico, compuesto por células enteras de la cepa Pdp11, se cultiva en TSAs durante 24 h a 22ºC. La preparación de una suspensión del probiótico, preferentemente sin proceso previo de liofilización o de inactivación física o química, se realiza mediante su incorporación en una matriz de alginato, preferentemente alginato sádico al 0,5%. La suspensión preparada se puede conservar sin pérdida significativa de viabilidad durante 20 - 30 días a 4ºC. Los productos alimenticios para peces son preparados mediante la adición en agitación de la suspensión de probiótico y, adicionalmente, CaCl2 50 mM, preferentemente mediante atomización.Universidad de Málaga (50%). Universidad de Almería (50%

Preparado de bacterias probióticas para su administración oral a peces cultivados basado en la encapsulación en hidrogeles de alginato

Número de solicitud: 201100469La invención consiste en un preparado para la administración oral del probiótico Shewanella PDP11 a peces. El preparado se caracteriza por contener células bacterianas viables de la cepa CECT 7627 encapsuladas en un hidrogel basado en alginato cálcico en forma de partículas esféricas de morfología uniforme y tamaño modificable para adaptarse a peces de distinto tamaño. Las cápsulas contienen aditivos organolépticos y/o nutricionales que actúan como atrayentes para los animales. El preparado mantiene viables a las bacterias durante periodos prolongados, es estable en medios acuáticos, y soporta el paso por el tubo digestivo de los peces, sobre los que ejerce efectos biológicos favorables. Su administración por vía oral es independiente del alimento habitual, evitando así la inactivación del microorganismo durante la fabricación del pienso.Universidad de Almerí

Gut microbiome specific changes in different behavioral profiles in a mouse social defeat stress model.

Comunicación de tipo PósterThe gut microbiome has arisen as one important modulator of general health, including brain function. In fact, disturbances in brain health are commonly mirrored in the microbiome, which could be contributing to pathology. One of the most common brain disorders is depression, which is tightly linked to environmental factors such as stress and drives alterations in regular behavior. However, not much is known about the role of the gut microbiome in response to stress and its relationship to behavior. In this study, the social defeat stress (SDS) paradigm was used as a depressive-like symptoms inducer in 8 w.o. male C57BL/6J mice for 10 days. Mice were segregated in stress resilient and sensitive according to behavior using K-means clustering and behavioral data was interpreted using principal component analysis. Then, the mice microbiome was extracted from fecal pellets after the stress protocol. DNA was extracted and purified followed by 16S (V3-V4) region amplification for sequencing. These data were analyzed to obtain diversity indexes and identify bacterial taxa within samples and groups. Data revealed that mice responded differently to the same stressor. Half the mice were found to have mild depressive-like symptoms whereas the other half showed profound alterations. Behavioral data was found to be explained in three factors: anhedonia, exploration, and motility. Stressed mice showed overall differences in their microbiome, being less diverse and populations associated with higher inflammation. Moreover, the healthy gut associated Verrucomicrobiae class was only identified in stress resilient mice, suggesting a possible relationship with their behavioral phenotype. Altogether, these results show a different behavioral response to stress in animals that reflects in their microbiome, which could be a key factor in determining stress resilience.This study was supported by Ministerio de Ciencia e Innovación - Plan Nacional I+D+I from Spain (PID2020-117464RB-I00) to CP and MP-M; FEDER/Junta de Andalucía - Proyectos I+D+I en el marco del Programa Operativo FEDER Andalucía 2014-2020 (UMA20-FEDERJA-112) to CP and MP-M; Consejería de Conocimiento, Investigación y Universidades, Junta de Andalucía (P20_00460) to CP. PC-P has been funded by the research project PID2020-117464RB-I00. Ministerio de Educación, Cultura y Deporte from Spain (FPU19/03629 to Infantes-López MI). Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Shewanella putrefaciens Pdp11 extracts protect against betanodavirus infection.

Viral nervous necrosis is caused by the nervous necrosis virus (Betanodavirus genus), a naked virus with two positive-sense RNA segments. Betanodaviruses are classified into four species, being RGNNV predominant in the Mediterranean area. RGNNV causes high mortality in several fish species, including European seabass. There are two vaccines to protect seabass against RGNNV infection. In this regard, the development of strategies to protect different fish species against different viruses, such as the use of probiotics, is a key issue for the aquaculture industry. Shewanella putrefaciens Pdp11, SpPdp11, is a fish probiotic with proven positive effects on seabream and Senegalese sole, protecting those species against bacterial pathogens; however, its antiviral activity is unknown. This study evaluates the anti-RGNNV activity of sonicated-SpPdp11 extracts in vitro and in vivo. The in vitro evaluation was performed on E11 cells following 3 assays: (i) neutralization, (ii) 6-h pre-adsorption, (iii) post-adsorption, determining the inhibition of RGNNV-induced CPEs and quantifying viral replication. The immunostimulatory activity of SpPdp11 extracts was also examined, analysing the transcription of mx, hsp70, tnfα, e3 and tlr3 in E11 cells. For the in vivo evaluation, two European seabass groups were established: (i) control group, (ii) experimental group, fed with commercial pellet supplemented with SpPdp11 extracts. Animals were fed for 30 days and subsequently challenged by intramuscular injection. SpPdp11 extracts compromised RGNNV replication in E11 cells (67.3% and 55% CPE inhibition in pre-adsorption and post-adsorption assays, respectively), and modulated the transcription of all the immune-related genes examined. The highest induction was obtained for mx gene. Regarding the in vivo results, 82% of fish fed with the SpPdp11-supplemented diet survived to RGNNV infection, whereas the survival rate of fish fed with the control diet was 64%.P18-RT-1067 (Proyecto de Excelencia, Junta de Andalucia), PID2020-113637RB-C22 (MINECO/AEI/FEDER, UE). Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Inhibition of nervous necrosis virus replication by Shewanella putrefaciens Pdp11 extract

One of the most relevant infectious diseases affecting farmed fish is the viral encephalopathy and retinopathy, caused by the nervous necrosis virus (NNV). The two commercial vaccines available against this virus have been designed to protect European seabass against the infection with a particular NNV species, RGNNV. Therefore, the development of strategies to protect different fish species against different viruses is a key issue for the aquaculture industry. In this regard, there are numerous studies focused on the use of probiotics, although their antiviral activity has been poorly investigated. Shewanella putrefaciens Pdp11, SpPdp11, is a probiotic with proven positive effects on gilthead seabream and Senegalese sole, protecting those species against several bac- terial pathogens; however, its antiviral activity remains to be investigated. The current study is a step forward in the use of probiotics against viral infections, evaluating the anti-RGNNV activity of sonicated-SpPdp11 extracts both, in vitro and in vivo. According to our results, SpPdp11 extracts compromised RGNNV multiplication in E11 cells, affecting viral assembly and/or exit rather than genome replication. Furthermore, this antiviral activity may be produced by the capacity of the SpPdp11-extract to induce an immune response in treated cells. This antiRGNNV activity has been confirmed in vivo, since 82% of fish fed with the SpPdp11-supplemented diet survived an experimental-RGNNV infection, whereas the survival rate of fish fed with the control diet was 64%. These results suggest that SpPdp11-supplemented feeding can be a promising prophylactic tool against RGNNV and encourage further research on other fish species and viral pathogens.This study has been supported by the projects P18-RT-1067 (Proyecto de Excelencia, Junta de Andalucia, Regional Government) and PID2020-113637RB-C22 (MINECO/AEI/FEDER, UE) (Spanish Government). Daniel Álvarez-Torres has been granted by the project PTA2020-018984-I. Funding for open access charge: Universidad de Málaga/CBUA

Identificación del probiótico Shewanella putrefaciens Pdp11 mediante PCR a través del transposón único que interrumpe al gen que codifica para la proteína fenazina.

Shewanella putrefaciens Pdp11 es una cepa descrita como probiótico en peces de importancia acuícola. La secuenciación de su genoma ha permitido establecer comparaciones a nivel genómico con otras cepas patógenas pertenecientes al mismo género. Como parte del estudio del genoma de Pdp11 se han identificado la presencia de 6 transposones y su ausencia en 7 cepas de Shewanella.sp (Pérez Gómez Olivia et al., 2021). En este trabajo se plantea el uso del transposón que interrumpe la proteína PhzE, implicada en la biosíntesis de la fenazina, para la identificación especifica de SpPdp11. Así como, la puesta a punto de la PCR para determinar la sensibilidad de los cebadores en la identificación del probiótico. En los cultivos acuícola, Shewanella putrefaciens Pdp11 se administra a los peces mediante su dieta en concentraciones de 10^9 ufc/gr de pienso, este trabajo permitiría la futura identificación y cuantificación del probiótico en muestras intestinales, así como el estudio del potencial de colonización del mismo.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Assessment of in vitro postbiotic capabilities of the probiotic Shewanella putrefaciens pdp11 growing under different cultivation conditions containing microalgae dietary supplements widely used in aquaculture.

Probiotics have been established as potential tools to improve host health and environmental quality in aquaculture. For example,Shewanella putrefaciens Pdp11 (SpPdp11), a strain described as a probiotic for use in aquaculture (reviewedby Cámara et al. 2020). Despite the proven health benefits of probiotics, recent evidence suggests that bacterial viability is not necessary to attain effects. Thus, postbiotics have emerged as potential opportunities in the field of functional foods. The International Scientific Association for Probiotics and Prebiotics (ISAPP) convened a panel that defined postbiotics as a “preparation of inanimate microorganisms and/or their components that confer a health benefit to the host”. The present work evaluates the potential postbiotic, as extracellular products (ECPs), of SpPdp11 grown undern different cultivation conditions. These included different culture media composed of a blend of microalgae, to observe a possible synergistic effect. The ECPs obtained were evaluated for their cytotoxicity against different fish cell lines, enzymatic and antibacterial activity, and their effect on biofilm formation by several fish pathogensproyecto PID2020-113637RB-C22 de investigación financiado por MCIN/AEI/10.13039/ 50110001103

Evaluación de la influencia de diferentes condiciones de cultivo en el potencial postbiótico de productos extracelulares de Shewanella putrefaciens Pdp11

Shewanella putrefaciens Pdp11 es una cepa probiótica que ejerce importantes efectos beneficiosos en peces cultivados. Sin embargo, no hay conocimiento sobre su potencial postbiótico en la industria acuícola. Dado que la producción de metabolitos postbióticos puede verse afectada por diferentes factores, el presente trabajo estudia los efectos en diferentes condiciones de crecimiento: (i) medios de cultivo, (ii) temperatura, (iii) tiempo de incubación y (iv) salinidad sobre el potencial postbiótico de los productos extracelulares (EPCs) de Pdp11. Para ello, se han analizado sus capacidades hidrolíticas, antibacterianas, antivirales y citotóxicas. Los resultados obtenidos han demostrado que las potenciales características postbióticas de Pdp11 dependen de las condiciones de crecimiento estudiadas, mostrando diferentes perfiles de hidrólisis enzimática, un efecto antivírico en el título del Virus de la Necrosis Nerviosa (NNV), un efecto estimulante en la viabilidad de diferentes líneas celulares, y un efecto en la formación de biopelícula de patógenos de peces.Este trabajo ha sido financiado por MINECO (#PID2020-113637RB-C22). Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tec

Potencial postbiótico de productos extracelulares de bacterias probióticas aisladas del tracto gastrointestinal de dorada (Sparus aurata)

Los probióticos en acuicultura son conocidos por sus efectos beneficiosos sobre el tracto digestivo y el sistema inmune en peces cultivados. Sin embargo, debido a los inconvenientes que genera incluir un microorganismo vivo en el proceso de producción de alimentos, derivados de los probióticos están cobrando cada vez más interés, como el caso de los postbióticos. A partir del aislamiento y caracterización de potenciales bacterias autóctonas del intestino de especímenes de Sparus aurata alimentadas con una dieta rica en algas, se han seleccionado 4 cepas bacterianas: UMA 140, UMA 143, UMA 169 y UMA 216. El presente trabajo evalúa el potencial postbiótico de estas cepas sometidas a diferentes condiciones de cultivos enriquecidos con algas, analizando su capacidades hidrolíticas, antibacterianas y citotóxicas con el objetivo de seleccionar el mejor o los mejores postbióticos para incluir en alimentos acuícolas.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tec