Subunit vaccine candidates against Aeromonas salmonicida in rainbow trout Oncorhynchus mykiss

Aeromonas salmonicida subsp. salmonicida is the etiological agent of furunculosis and a major fish health problem in salmonid aquaculture worldwide. Injection vaccination with commercial mineral oil-adjuvanted bacterin vaccines has been partly successful in preventing the disease but in Danish rainbow trout (Oncorhynchus mykiss, Walbaum) aquaculture furunculosis outbreaks still occur. In this study we tested the efficacy of experimental subunit vaccines against A. salmonicida infection in rainbow trout. We utilized in silico screening of the proteome of A. salmonicida subsp. salmonicida strain A449 and identified potential protective protein antigens that were tested by in vivo challenge trial. A total of 14 proteins were recombinantly expressed in Escherichia coli and prepared in 3 different subunit vaccine combinations to immunize 3 groups of rainbow trout by intraperitoneal (i.p.) injection. The fish were exposed to virulent A. salmonicida 7 weeks after immunization. To assess the efficacy of the subunit vaccines we evaluated the immune response in fish after immunization and challenge infection by measuring the antibody levels and monitoring the survival of fish in different groups. The survival of fish at 3 weeks after challenge infection showed that all 3 groups of fish immunized with 3 different protein combinations exhibited significantly lower mortalities (17-30%) compared to the control groups (48% and 56%). The ELISA results revealed significantly elevated antibody levels in fish against several protein antigens, which in some cases were positively correlated to the survival

A benchmarking model for Maintenance and Support assignments in an ERP environment

We present a benchmarking model on how to track and eliminate dissimilarities in M&S assignments, supporting different Enterprise Resource Planning (ERP) environments. ERP-user organizations, SAP systems, infrastructure and level of service quality are some examples of elements that will drive M&S effort and cost. In each of these aspects, there are cost drivers that influence the M&S process in different extents. Our model identifies the impact of the many different parameters for the uncovering of Maintenance and Support (M&S) costs and benefits within, or between, organizations. To be able to compare parameters that affect the M&S work load we have developed a complexity calculator, which work to normalize such parameters. The complexity calculator compares the parameters of interest with a mean-assignment calculated from our reference database, which will create a complexity index and, thus, enables the comparison of all factors that may be of consideration. After receiving the index, we suggest a comparison with one or several peer-groups to further remove variations between the assignments, for instance organizations with similar number of users. Furthermore, our model enables the possibility of evaluation and measurement of assignments cost, benefits, system characteristics and M&S staff productivity with the utilization of key performance indicators (KPI) on both qualitative and quantitative measures. Additionally, the model provides functionality for cost and effort prognostication. In conclusion, we show how to compare M&S assignments on several aspects, which facilitate cost and service strategies for M&S organizations. The ERP using organizations will acquire an increased awareness of M&S cost drivers, which facilitates development of cost efficient strategies. Thus, our model serves as a good ground in benchmarking assignments

In silico prediction of <i>Gallibacterium anatis</i> pan-immunogens

The Gram-negative bacterium Gallibacterium anatis is a major cause of salpingitis and peritonitis in commercial egg-layers, leading to reduced egg production and increased mortality. Unfortunately, widespread multidrug resistance and antigenic diversity makes it difficult to control infections and novel prevention strategies are urgently needed. In this study, a pan-genomic reverse vaccinology (RV) approach was used to identify potential vaccine candidates. Firstly, the genomes of 10 selected Gallibacterium strains were analyzed and proteins selected on the following criteria; predicted surface-exposure or secretion, none or one transmembrane helix (TMH), and presence in six or more of the 10 genomes. In total, 42 proteins were selected. The genes encoding 27 of these proteins were successfully cloned in Escherichia coli and the proteins expressed and purified. To reduce the number of vaccine candidates for in vivo testing, each of the purified recombinant proteins was screened by ELISA for their ability to elicit a significant serological response with serum from chickens that had been infected with G. anatis. Additionally, an in silico prediction of the protective potential was carried out based on a protein property prediction method. Of the 27 proteins, two novel putative immunogens were identified; Gab_1309 and Gab_2312. Moreover, three previously characterized virulence factors; GtxA, FlfA and Gab_2156, were identified. Thus, by combining the pan-genomic RV approach with subsequent in vitro and in silico screening, we have narrowed down the pan-proteome of G. anatis to five vaccine candidates. Importantly, preliminary immunization trials indicated an in vivo protective potential of GtxA-N, FlfA and Gab_1309

Experimental setup presenting the different groups, sample size, mortality and relative percentage of survival (RPS).

<p>Experimental setup presenting the different groups, sample size, mortality and relative percentage of survival (RPS).</p

Correlation between mean antibody (diluted 1:100) levels for individual proteins at 7 weeks post-immunization (wpi) and survival in different experimental groups at 24 days post-challenge (dpc).

<p>Antibody levels were measured from 10 fish per group (5 fish per duplicate tank) by ELISA one day before challenge.</p

Recombinant construct design, protein type and vaccine formulation.

<p>Recombinant construct design, protein type and vaccine formulation.</p

Levels of antibodies in serum of fish against individual VacC vaccine proteins.

<p>Sera (diluted 1:100) was measured by ELISA from 10 fish per group (5 fish per duplicate tank) at 7 weeks post-immunization (wpi) (a) and at 3 weeks post-challenge (wpc) (b). Asterisks (*) represent p values (* = P<0.05, ** = P< 0.01, *** = P< 0.001) when compared to saline control group.</p

Levels of <i>A</i>. <i>salmonicida</i>-specific antibodies in serum measured by ELISA at 7 weeks post-immunization (wpi) and 3 weeks post- challenge (wpc).

<p>Sera (diluted 1:500) were analyzed from 10 fish per group (5 fish per duplicate tank). Asterisks (*) represent p values (*p<0.05, **p< 0.01, ***p< 0.001) compared to saline control group.</p