Possible Mechanisms of Action of Two Pseudomonas fluorescens Isolates as Probiotics on Saprolegniosis Control in Rainbow Trout (Oncorhynchus mykiss Walbaum)

© 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).[EN] Probiotics have been proposed as one of the alternatives to the chemical treatments currently used in aquaculture. Recently, the possible usefulness of certain microorganisms, mainly bacteria, has been highlighted as a potential biocontrol for saprolegniosis. In the present work we investigated the possible mechanisms of action of two isolates of Pseudomonas fluorescens (LE89 and LE141) with proven ability to reduce Saprolegnia parasitica infection in rainbow trout under experimental conditions when they are added to the tank water. The stimulation of the innate immune response and the production of siderophores and bioactive substances inhibiting S. parasitica present in cells and supernatants of LE89 and LE141 were studied. Regarding the immune response the only noteworthy points were the increase in the phagocytic activity of macrophages and the concentration of serum proteins when LE141 was administered. Both bacteria produced siderophores. When analyzing the protein substances present in supernatants, it was observed that in both isolates the proteins with inhibitory activity present might be siderophores. In LE141, besides siderophores, a protein of 66 kDa was identified in the fraction responsible for inhibition. To sum up, the two P. fluorescens isolates might be usable for biocontrol of saprolegniosis and that the mode of action of these bacteria is likely to be related to the production of siderophores.SIThis research was funded by “Ministerio de Ciencia e Innovación” (AGL2010-18562) and co-financed through the European Regional Development Fund (ERDF). C.G.P. was awarded a predoctoral contract by the “Consejería de Educación” of the regional government “Junta de Castilla y León”, co-financed through the European Social Fund.The authors are grateful for the technical assistance provided by Gloria Fernández-Bayón, Leandro Rodríguez Aparicio, and Maria Camino López Fierro of the University of León. The fish farm Los Leoneses, in León, supplied the rainbow trout

Mixed germ cell-stromal testicular neoplasia in a crucian carp (Carassius carassius L.): a case report

p. 114-119This report documents a case of spontaneously occurring gonadal neoplasm in a crucian carp (Carassius carassius L.) caught in the wild. On the basis of the histopathological findings, the neoplasm was diagnosed as a mixed germ cell-stromal testicular neoplasia (gonadoblastoma).S

Limited Probiotic Effect of Enterococcus gallinarum L1, Vagococcus fluvialis L21 and Lactobacillus plantarum CLFP3 to Protect Rainbow Trout against Saprolegniosis

[EN] Previous studies have demonstrated that the strains Enterococcus gallinarum L1, Vagococcus fluvialis L21 and Lactobacillus plantarum CLFP3 are probiotics against vibriosis or lactococosis in sea bass or rainbow trout. In this study, the utility of these bacterial strains in the control of saprolegniosis was evaluated. For this purpose, both in vitro inhibition studies and competition for binding sites against Saprolegnia parasitica and in vivo tests with experimentally infected rainbow trout were carried out. In the in vitro tests, the three isolates showed inhibitory activity upon mycelium growth and cyst germination and reduced the adhesion of cysts to cutaneous mucus; however, this effect depended on the number of bacteria used and the incubation time. In the in vivo test, the bacteria were administered orally at 108 CFU g−1 in the feed or at 106 CFU ml−1 in the tank water for 14 days. None of the three bacteria showed protection against S. parasitica infection either through water or feed, and the cumulative mortality reached 100% within 14 days post infection. The obtained results show that the use of an effective probiotic against a certain disease in a host may not be effective against another pathogen or in another host and that the results obtained in vitro may not always predict the effects when used in vivo.SIWe are grateful for the assistance provided by the laboratory technician Gloria Fernández–Bayón; the fish farm Los Leoneses in León, which supplied the rainbow trout; the “Servicio Territorial de Medio Ambiente de León” of the “Junta de Castilla y León”, which supplied the brown trout; and Skretting España S.A., which donated some of the feed.This study was funded through research project AGL2014-54683-R of the Spanish “Ministerio de Economía y Competitividad” and cofinanced through the European Regional Development Fund (ERDF). C.G.P. was awarded a predoctoral contract by the “Consejería de Educación” of the regional government “Junta de Castilla y León” cofinanced through the European Social Fund

Clinical evaluation and antibody responses in sheep after primary and secondary experimental challenges with the mange mite Sarcoptes scabiei var. ovis

P. 109-116In this work the clinical evolution and the specific serum IgG and IgE antibody responses in sheep after primary (n = 10) and secondary (n = 4) experimental challenges with the mange mite Sarcoptes scabiei var. ovis were studied. The primary infection was characterized by the development of mange lesions in all sheep, a detection of live S. scabiei mites in 70% skin scrapings taken in week 10 post-challenge (PC), strongly raised and sustained specific IgG levels and a more moderate but continuous rise in specific IgE levels. Seroconversion was detected for IgG and IgE by ELISA in 90% and 60% of the sheep in week 8 PC, respectively. By Western-blotting (WB), ten IgG-reactive bands (36–120 kDa) and four IgE-reactive bands (90–180 kDa) were observed in week 8 PC. Following the secondary challenge the ewes developed a smaller area of mange lesion than that seen following primary challenge and live S. scabiei mites were not detected in skin scrapings collected in week 8 PC, suggesting that sheep had developed immunity to re-infection. Compared to primary infection, the specific IgG secondary antibody levels were transient, but in contrast there was an anamnestic IgE response, resulting in an elicitation of specific serum IgE levels in week 2 PC significantly higher than those demonstrated after primary infection. WB analysis revealed one additional IgG-reactive band (180 kDa) and no additional IgE-reactive bands. Determining the immunodiagnostic or vaccination value of the IgG-reactive antigens and IgE-reactive allergens detected requires further studies.S

Prevalence of serum antibodies against Saprolegnia parasitica in wild and farmed brown trout Salmo trutta

p. 17-22The prevalence of serum antibodies against Saprolegnia parasitica in wild and farmed brown trout Salmo trutta from the province of León (NW Spain) was studied by enzyme-linked immunosorbent assay (ELISA). Blood samples from healthy and Saprolegnia-infected brown trout were collected over 2 yr with a seasonal periodicity (January, April, July and October) from a hatchery and river with frequent presence of saprolegniosis (River Porma) and from a river in which the disease was rarely observed (River Omaña). The individual prevalence was 30.1%, but statistically significant differences were observed between the prevalence in trout from the hatchery (43.0%), from River Porma (31.8%) and from River Omaña (6.4%) and also between the prevalence observed in October (42.9%) and the values obtained in January (24.8%), April (22.7%) and July (27.5%). There was no difference between the seroprevalence in females (34.8%) and males (38.2%), but a positive correlation between raised serum antibody levels and larger (older) fish was found. The low prevalence of antibodies observed in Saprolegnia-infected trout (18.0%) suggests possible immune suppression and the lack of an effective specific immune response in fish with saprolegniosis.S

Sobre un foco de tiña en cabras por Trichophyton Verrucosum

[ES] Sobre un foco de tiña en cabras por Trichophyton Verrucosu

Sobre unos casos de infección humana por Trichophyton mentagrophytes transmitido por ratones de laboratorio.

[ES] Sobre unos casos de infección humana por Trichophyton mentagrophytes transmitido por ratones de laboratorio

Adhesion to brown trout skin mucus, antagonism against cyst adhesion and pathogenicity to rainbow trout of some inhibitory bacteria against Saprolegnia parasitica

P. 35-44Biological control of saprolegniosis with bacteria might be an alternative to the use of chemical compounds. Among criteria for the selection of such bacteria are their absence of pathogenicity to fish and their ability to prevent adhesion of the pathogen to the skin mucus. The pathogenicity to rainbow trout of 21 bacterial isolates with in vitro inhibitory activity against Saprolegnia parasitica was studied. Fifteen of the isolates, identified as Aeromonas sobria, Pantoea agglomerans, Pseudomonas fluorescens, Serratia fonticola, Xanthomonas retroflexus and Yersinia kristensenii, were non-pathogenic when injected into rainbow trout. Their capacity to adhere to the skin mucus of male and female brown trout and to reduce the adhesion of S. parasitica cysts under exclusion, competition and displacement conditions was tested. The 15 bacterial isolates showed a low adhesion rate, ranging between 1.7% (for an A. sobria isolate) and 15.3% (a P. fluorescens isolate). This adhesion was greater in the case of mucus from male brown trout than from females. Similarities in the adhesion to male mucus and other substrates and correlation to that observed to polystyrene suggest that adhesion to skin mucus does not depend on the substrate. A high percentage (88.9%) of the S. parasitica cysts adhered to the skin mucus of male brown trout. Almost all of the bacteria reduced this adhesion ratio significantly under exclusion and competition conditions. However, only half of the isolates displaced cysts from skin mucus, and more bacterial cells were necessary for this effect. A novel method to study the adhesion of S. parasitica cysts to skin mucus of trout and their interactions with inhibitory bacteria is described.S

Antibody response of brown trout Salmo trutta injected with pathogenic Saprolegnia parasitica antigenic extracts

P. 107-111Brown trout Salmo trutta injected with antigenic extracts from a pathogenic isolate of Saprolegnia parasitica developed specific antibodies that were detected by enzyme-linked immunosorbent assay (ELISA), immunofluorescence (IF) and Western blotting (WB), but not by immunodiffusion (ID). Three groups of five 2 yr old brown trout were injected intraperitoneally with 3 different antigenic extracts: small hyphal fragments (HF) and soluble extracts from sonicated mycelia grown in medium with or without β-sytosterol (SEB and SE, respectively). In the 2 groups injected with SE and SEB, antibodies were found in 66.7% of the serum samples by ELISA, 54.5% by IF and 48.5% by WB. In the group injected with HF, only 1 trout survived the experiment, and in this fish only 1 sample was positive by ELISA. The results obtained by ELISA and IF were similar and show that there is cross-reaction between the antigens used. By WB, the proteins most frequently recognised were 2 proteins of 25 and 29 kDa. No significant differences were found in the groups injected with SE or SEB.S