Piscirickettsiosis or salmonid rickettsial septicaemia (SRS) caused by Piscirickettsia salmonis constitutes one of the main problems in farmed salmonid and marine fishes. Since the first reports of the disease, it has been successfully isolated and maintained in eukaryotic cellculture systems, but these systems are time-consuming, the media are costly, and eliminating heavily contaminated host cell debris is difficult. In this report, we describe a marine-based broth supplemented with L-cysteine, named AUSTRAL-SRS broth, that facilitates superior growth of P. salmonis strains. Strains reached an optical density of approximately 1.8 when absorbance was measured at 600 nm after 6 d incubation at 18°C. Several passages (n = 6) did not alter the culture kinetics. We report for the first time the purification of DNA, lipopolysaccharide (LPS) and whole membrane protein obtained from P. salmonis grown in this liquid medium, and thus provide a suitable platform to simplify the preparation of P. salmonis cells for genetic and serological studies. Moreover, the results of the cytopathic effect test showed that P. salmonis grown in AUSTRAL-SRS broth maintained their virulence properties, inducing apoptosis after 3 d. This makes the medium a good candidate for the successful growth of P. salmonis and an excellent basis for the development of low cost vaccines. KEY WORDS: Piscirickettsia salmonis · Broth · CultureResale or republication not permitted without written consent of the publisher

Three different lyric bacteriophages (BPs) were isolated from the sewage system of commercial chicken flocks and used to reduce Salmonella Enteritidis (SE) colonization from experimental chickens. Ten-day-old chickens were challenged with 9.6 x 10(5) colony-forming units (CFU)/ml of a SE strain and treated by coarse spray or drinking water with a cocktail of the three phages at a multiplicity of infection (MO1) of 10(3) plaque-forming units (PFU) 24 hr prior to SE challenge. Chickens were euthanatized at day 20 of age for individual SE detection, quantitative bacteriology, and phage isolation from the intestine and from a pool of organs. SE detection was performed by both bacteriologic culture and genome detection by polymerase chain reaction (PCR). Qualitative bacteriology showed that aerosol-spray delivery of BPs significantly reduced the incidence of SE infection in the chicken group (P = 0.0084) to 72.7% as compared with the control group (100%). In addition, SE counts showed that phage delivery both by coarse spray and drinking water reduced the intestinal SE colonization (P < 0.01; P < 0.05, respectively). BPs were isolated at 10 days postinfection from the intestine and from pools of organs from BP-treated chickens. We conclude that the phage treatment, either by aerosol spray or drinking water, may be a plausible alternative to antibiotics for the reduction of Salmonella infection in poultry.

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Pseudomonas syringae pv. actinidiae (Psa) is the causal agent of a bacterial canker in kiwifruit plants and has caused economic losses worldwide. Currently, the primary strategies to control this pathogen include the use of copper-based compounds and even antibiotics. However, the emergence of isolates of Psa that are resistant to these agrochemicals has raised the need for new alternatives to control this pathogen. Bacteriophages have been proposed as an alternative to control bacterial infections in agriculture, including Psa. Here, we show the isolation and characterization of 13 phages with the potential to control Psa infections in kiwifruit plants. The phages were characterized according to their host range and restriction fragment length polymorphism (RFLP) pattern. Four phages were selected according to their lytic effect on the bacteria and their tolerance to different environmental conditions of pH (4–7), temperature (4–37 °C), and solar radiation exposure (30 and 60 min). The selected phages (CHF1, CHF7, CHF19, and CHF21) were sequenced, revealing a high identity with the podophage of Psa phiPSA2. In vitro assays with kiwifruit leaf samples demonstrated that the mixture of phages reduced the Psa bacterial load within three hours post-application and was able to reduce the damage index in 50% of cases. Similarly, assays with kiwifruit plants maintained in greenhouse conditions showed that these phages were able to reduce the Psa bacterial load in more than 50% of cases and produced a significant decrease in the damage index of treated plants after 30 days. Finally, none of the selected phages were able to infect the other bacteria present in the natural microbiota of kiwifruit plants. These results show that bacteriophages are an attractive alternative to control Psa infections in kiwifruit plants.

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A combination of three different Salmonella-specific bacteriophages (BPs) and one competitive exclusion (CE) product were used to reduce Salmonella Enteritidis (SE) colonization in experimentally infected chickens. Equal numbers of 7-day-old chickens were used in each of three groups: a CE group (treated with CE), a BP group (treated with BP), and a CE-plus-BP group (treated with both products). The CE product was administered via coarse spray at 1 day of age and the cocktail of three BPs was given via spray at 6 days of age using a multiplicity of infection of 10(3) plaque-forming units. All the experimental groups, except a healthy control group, were challenged orally with 2.95 x 10(5) colony-forming units (CFU)/ml of an SE strain at 7 days of age. Seven days postchallenge, the chickens were euthanatized for individual SE detection, quantitative bacteriology, and phage isolation from ceca and an internal organ pool. The qualitative bacteriology demonstrated that the use of the CE product diminished the incidence of SE to 75.7% and the mixture of BPs reduced it to 80%; when CE plus BP were used, the incidence dropped significantly to 38.7% (P < 0.0001), as compared with the infection control group (100%). A significant difference in the incidence was observed between the CE and the CE-plus-BP groups, and the BP and the CE-plus-BP groups (P = 0.0027 and P = 0.0010, respectively). The mean SE cecal count diminished with the use of CE plus BP (1.6 x 10(2) CFU/g, P = 0.0003) compared with the control group (1.56 x 10(5) CFU/g), the CE group (4.23 x 10(3) CFU/g), and the BP group (9.48 x 10(3) CFU/g). On the basis of the present study, it may be concluded that the use of both types of biocontrollers can be an effective method for reducing SE colonization in commercial chickens, but further basic and applied research is needed.

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