Protective immunity in rainbow trout Oncorhynchus mykiss following immunization with distinct molecular mass fractions isolated from Flavobacterium psychrophilum

LaFrentz, Benjamin R.; LaPatra, Scott E.; Jones, G. R.; Cain, Kenneth D.

doi:10.3354/dao059017

Cited by 73 publications

(101 citation statements)

References 36 publications

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“…Daily mortalities were recorded, and at least 25% of dead fish per tank were evaluated for signs of BCWD and isolation of yellow-pigmented bacteria by culturing of the liver, spleen, and kidneys on TYES agar at 15°C for at least 96 h. The relative percent survival (RPS) was calculated according to the method of LaFrentz et al (37), using the following formula: RPS ϭ [1 Ϫ (cumulative percent mortality of treatment group/cumulative percent mortality of positive-control group)] ϫ 100. Animal challenge experiments conformed to a protocol approved by the University of Idaho Institutional Animal Care and Use Committee.…”

Section: Methodsmentioning

confidence: 99%

Entericidin Is Required for a Probiotic Treatment (Enterobacter sp. Strain C6-6) To Protect Trout from Cold-Water Disease Challenge

Schubiger

Orfe

Sudheesh

et al. 2015

Appl Environ Microbiol

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c Flavobacterium psychrophilum causes bacterial cold-water disease in multiple fish species, including salmonids. An autochthonous Enterobacter strain (C6-6) inhibits the in vitro growth of F. psychrophilum, and when ingested as a putative probiotic, it provides protection against injection challenge with F. psychrophilum in rainbow trout. In this study, low-molecular-mass (<3 kDa) fractions from both Enterobacter C6-6 and Escherichia coli K-12 culture supernatants inhibited the growth of F. psychrophilum. The <3-kDa fraction from Enterobacter C6-6 was analyzed by SDS-PAGE, and subsequent tandem mass spectroscopy identified EcnB, which is a small membrane lipoprotein that is a putative pore-forming toxin. Agar plate diffusion assays demonstrated that ecnAB knockout strains of both Enterobacter C6-6 and E. coli K-12 no longer inhibited F. psychrophilum (P < 0.001), while ecnAB-complemented knockout strains recovered the inhibitory phenotype (P < 0.001). In fish experiments, the engineered strains (C6-6 ⌬ecnAB and C6-6 ⌬ecnAB) and the wild-type strain (C6-6) were added to the fish diet every day for 38 days. On day 11, the fish were challenged by injection with a virulent strain of F. psychrophilum (CSF 259-93). Fish that were fed C6-6 had significantly longer survival than fish fed the ecnAB knockout strain (P < 0.0001), while fish fed the complemented knockout strain recovered the probiotic phenotype (P ‫؍‬ 0.61). This entericidin is responsible for the probiotic activity of Enterobacter C6-6, and it may present new opportunities for therapeutic and prophylactic treatments against similarly susceptible pathogens. P robiotics are defined as live or dead microbial feed supplements that provide health benefits to the host animal (1). Probiotic strains of Bifidobacterium, Lactobacillus, and Streptococcus (2) have been used successfully in people and terrestrial animals, including poultry (3) and cattle (4), for the past century. Probiotic supplementation has gained attention from the aquaculture industry during the last 2 decades, especially for shrimp production, where antibiotic treatment has been reduced dramatically in favor of supplementing feed with commercially available probiotics composed of Bacillus subtilis and Vibrio alginolyticus (5, 6). In aquaculture, a probiotic is further defined as a live microbial cell that, when administered via the feed or to the rearing water, benefits the host by improving disease resistance, health and growth performance, feed utilization, stress response, and meat quality (7,8). The mechanism of action of probiotics has been defined vaguely as improving the host's "microbial balance" (9) or the balance of the ambient environment, but molecular mechanisms are rarely investigated further, even though a number of applied studies have demonstrated probiotic efficacy against a variety of Gram-negative and Gram-positive pathogens in fish and shellfish (7).In the case of salmonid species, such as Atlantic salmon (Salmo salar Linnaeus) and rainbow trout (Oncorhynchus ...

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Section: Methodsmentioning

confidence: 99%

Entericidin Is Required for a Probiotic Treatment (Enterobacter sp. Strain C6-6) To Protect Trout from Cold-Water Disease Challenge

Schubiger

Orfe

Sudheesh

et al. 2015

Appl Environ Microbiol

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“…Crump et al (2001) identified several protein and carbohydrate antigens of F. psychrophilum and subsequently characterized and expressed a ~20 kDa hostrecognized protein antigen for use in a recombinant protein vaccine (Crump et al 2005). LaFrentz et al (2004) identified 3 immunogenic regions of the bacterium and demonstrated a high level of protection in rainbow trout immunized with antigens in the 70 to 100 kDa molecular mass range. Additionally, Rahman et al (2002) demonstrated that outer membrane components of the bacterium were highly protective.…”

Section: Introductionmentioning

confidence: 99%

Identification of potential vaccine target antigens by immunoproteomic analysis of a virulent and a non-virulent strain of the fish pathogen Flavobacterium psychrophilum

Sudheesh

LaFrentz²,

Call³

et al. 2007

Dis. Aquat. Org.

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“…Only seven strains were positive for all five putative virulence genes (data not shown). Three of the isolates used in this analysis were known to be highly virulent in a trout challenge model: CSF 259-93, SH3-81, and 950106-1/1 (13,15,18). Only virulence gene candidate csf1-d7 (BspA like) is present in all three of these isolates and thus may be a target for vaccine development and studies of pathogenic mechanisms.…”

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confidence: 99%

Combining Suppression Subtractive Hybridization and Microarrays To Map the Intraspecies Phylogeny of Flavobacterium psychrophilum

et al. 2005

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Reciprocal subtractive libraries were prepared for two strains of Flavobacterium psychrophilum, one virulent and the other avirulent in a trout challenge model. Unique clones were sequenced and their distribution assessed among 34 strains. The analysis showed that F. psychrophilum is composed of two genetic lineages, possibly reflecting host specificity.Flavobacterium psychrophilum is the causative agent of bacterial coldwater disease and rainbow trout fry syndrome, both of which affect salmonid fish and impact commercial aquaculture and resource enhancement hatcheries worldwide (12,19,21). Coho salmon (Oncorhynchus kisutch [Walbaum]) and rainbow trout (O. mykiss) are particularly susceptible, although F. psychrophilum also infects other fish species (21). F. psychrophilum strains vary greatly in the ability to establish disease (virulence). For example, one well-studied strain (ATCC 49418 [4,6]) is unable to cause significant mortality (9) while strain CSF 259-93 causes high mortality (16) in a trout challenge model. There are no commercial vaccines available for bacterial coldwater disease, although several research groups have active programs in this area (11,14,20). From these studies we know that not all strains elicit an antibody response that is effective against other strains and consequently there may be considerable genetic variation between strains.The goals of this project were (i) to examine genetic differences between two strains of F. psychrophilum and ATCC 49418 [16]) and (ii) to assess the extent and distribution of genetic variation between other strains relative to geographical source and host species. Strains for the latter assessment were chosen based on availability at the time of this study. All strains were stored at Ϫ80°C and were cultured at 16 to 17°C in tryptone yeast extract salts medium (TYES; 0.4% tryptone, 0.04% yeast extract, 0.05% calcium chloride, 0.05% magnesium sulfate, pH 7.2). Genomic DNA (gDNA) was extracted using the DNeasy tissue kit (QIAGEN, Valencia, CA).We prepared reciprocal suppression subtractive hybridization libraries for the two strains by using the PCR-Select Bacterial Genomic Subtraction Kit (Clontech, Palo Alto, CA) according to the manufacturer's protocol, except that tester and driver gDNAs were restriction enzyme digested with RsaI (supplied with the kit) and DraI (New England Biolabs, Inc,, Beverly, MA) and the hybridization step was at 59°C. Resulting DNA fragments were cloned in pCR2.1 (Invitrogen, Carlsbad, CA), and 576 randomly chosen recombinant clones were used to make a microarray as previously described (8, 10). gDNA (0.5 g) was nick translated for 2 h in the presence of biotindATP (BioNick Labeling System; Invitrogen) and hybridized to the microarray (data not shown). Microarray slides were processed, imaged, and analyzed as described previously (23). Using stringent selection criteria, where the median probe intensity was Ն95% of the maximum pixel intensity in one strain and Յ5% of the maximum pixel intensity in the other strain, we identified...

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Protective immunity in rainbow trout Oncorhynchus mykiss following immunization with distinct molecular mass fractions isolated from Flavobacterium psychrophilum

Cited by 73 publications

References 36 publications

Entericidin Is Required for a Probiotic Treatment (Enterobacter sp. Strain C6-6) To Protect Trout from Cold-Water Disease Challenge

Entericidin Is Required for a Probiotic Treatment (Enterobacter sp. Strain C6-6) To Protect Trout from Cold-Water Disease Challenge

Identification of potential vaccine target antigens by immunoproteomic analysis of a virulent and a non-virulent strain of the fish pathogen Flavobacterium psychrophilum

Combining Suppression Subtractive Hybridization and Microarrays To Map the Intraspecies Phylogeny of Flavobacterium psychrophilum

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