Development of a DIVA subunit vaccine against Actinobacillus pleuropneumoniae infection

Maas, Alexander; Meens, Jochen; Baltes, Nina; Hennig-Pauka, Isabel; Gerlach, Gerald-F.

doi:10.1016/j.vaccine.2006.06.047

Cited by 33 publications

(35 citation statements)

References 33 publications

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“…This subunit vaccine was postulated to be enriched in outer membrane lipoproteins as suggested by the presence of transferrin binding lipoprotein B (TbpB) and the outer membrane lipoprotein A (OmlA) whereas periplasmic, cytoplasmic and membrane proteins were absent [20]. Vaccination of pigs with this vaccine derived from A. pleuropneumoniae apxIIA -mutants of serotypes 1, 2 and 5 showed good protective efficacy upon homologous challenge with serotype 2 and cross-protection against serotype 9 and facilitated the differentiation between infected and vaccinated animals [21]. …”

Section: Introductionmentioning

confidence: 99%

Proteomic and immunoproteomic characterization of a DIVA subunit vaccine against Actinobacillus pleuropneumoniae

et al. 2011

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BackgroundProtection of pigs by vaccination against Actinobacillus pleuropneumoniae, the causative agent of porcine pleuropneumonia, is hampered by the presence of 15 different serotypes. A DIVA subunit vaccine comprised of detergent-released proteins from A. pleuropneumoniae serotypes 1, 2 and 5 has been developed and shown to protect pigs from clinical symptoms upon homologous and heterologous challenge. This vaccine has not been characterized in-depth so far. Thus we performed i) mass spectrometry in order to identify the exact protein content of the vaccine and ii) cross-serotype 2-D immunoblotting in order to discover cross-reactive antigens. By these approaches we expected to gain results enabling us to argue about the reasons for the efficacy of the analyzed vaccine.ResultsWe identified 75 different proteins in the vaccine. Using the PSORTb algorithm these proteins were classified according to their cellular localization. Highly enriched proteins are outer membrane-associated lipoproteins like OmlA and TbpB, integral outer membrane proteins like FrpB, TbpA, OmpA1, OmpA2, HgbA and OmpP2, and secreted Apx toxins. The subunit vaccine also contained large amounts of the ApxIVA toxin so far thought to be expressed only during infection. Applying two-dimensional difference gel electrophoresis (2-D DIGE) we showed different isoforms and variations in expression levels of several proteins among the strains used for vaccine production. For detection of cross-reactive antigens we used detergent released proteins of serotype 7. Sera of pigs vaccinated with the detergent-released proteins of serotypes 1, 2, and 5 detected seven different proteins of serotype 7, and convalescent sera of pigs surviving experimental infection with serotype 7 reacted with 13 different proteins of the detergent-released proteins of A. pleuropneumoniae serotypes 1, 2, and 5.ConclusionsA detergent extraction-based subunit vaccine of A. pleuropneumoniae was characterized by mass spectrometry. It contained a large variety of immunogenic and virulence associated proteins, among them the ApxIVA toxin. The identification of differences in expression as well as isoform variation between the serotypes implied the importance of combining proteins of different serotypes for vaccine generation. This finding was supported by immunoblotting showing the induction of cross-reactive antibodies against several surface associated proteins in immunized animals.

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Section: Introductionmentioning

confidence: 99%

Proteomic and immunoproteomic characterization of a DIVA subunit vaccine against Actinobacillus pleuropneumoniae

et al. 2011

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“…As clear differences in virulence between serotypes have been observed, the genetic basis should be investigated at the genomic level. Here, we present the draft genome sequences of the A. pleuropneumoniae serotypes 2 (strain 4226) and 6 (strain Femo).Previous studies of different serotypes of Actinobacillus pleuropneumoniae showed that there are significant variations among them at the DNA sequence level, supposed to cause differences in pathogenicity and immunogenicity (2,8). However, it is difficult to carry out more general studies of the immunity mechanisms of different serotypes, typing-based diagnosis, and multivalent genetically engineered vaccines due to the lack of complete genome sequences of the different serotypes (11).…”

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confidence: 99%

“…Previous studies of different serotypes of Actinobacillus pleuropneumoniae showed that there are significant variations among them at the DNA sequence level, supposed to cause differences in pathogenicity and immunogenicity (2,8). However, it is difficult to carry out more general studies of the immunity mechanisms of different serotypes, typing-based diagnosis, and multivalent genetically engineered vaccines due to the lack of complete genome sequences of the different serotypes (11).…”

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Draft Genome Sequences of Actinobacillus pleuropneumoniae Serotypes 2 and 6

et al. 2010

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Actinobacillus pleuropneumoniae is a bacterial pathogen that causes highly contagious respiratory infection in pigs and has a serious impact on the production economy and animal welfare. As clear differences in virulence between serotypes have been observed, the genetic basis should be investigated at the genomic level. Here, we present the draft genome sequences of the A. pleuropneumoniae serotypes 2 (strain 4226) and 6 (strain Femo).Previous studies of different serotypes of Actinobacillus pleuropneumoniae showed that there are significant variations among them at the DNA sequence level, supposed to cause differences in pathogenicity and immunogenicity (2,8). However, it is difficult to carry out more general studies of the immunity mechanisms of different serotypes, typing-based diagnosis, and multivalent genetically engineered vaccines due to the lack of complete genome sequences of the different serotypes (11).Draft genome sequences of serotypes 2 and 6 were assembled by combining Roche 454-FLX reads with Illumina Genome Analyzer IIx paired-end reads. The final assembly (CLCGenomicsWorkbench version 3.6, CLCbio) of the serotype 2 genome has a length of 2,314,315 bp (38 contigs), while the assembly of serotype 6 has a length of 2,375,501 bp (36 contigs). The average GC contents are 41.17% and 40.95% in serotypes 2 and 6, respectively, similar to those reported for other serotypes (1, 12). Using EasyGene (6, 9), 2,100 putative open reading frames were predicted for serotype 2 and 2,168 for serotype 6. Approximately 86% of the nucleotides were predicted to be involved in coding sequences, which is similar to results reported for other finished A. pleuropneumoniae genomes (1, 12). There are 86 and 136 genes found to be specific for serotypes 2 and 6, respectively, that do not have any homologue in other reported serotypes. rRNA genes were identified by RNAmmer (5). Serotype 2 harbors six rRNA operons (16S-23S-5S rRNA) and three additional 5S rRNA genes, while six rRNA operons and one additional 5S rRNA gene are present in serotype 6. Using the tRNAscan-SE server (7), 61 and 59 tRNA operons were predicted for serotypes 2 and 6, respectively.Since porcine pleuropneumonia caused by A. pleuropneumoniae leads to large economic losses for the swine industry (3), it is of considerable interest to investigate the virulence factors of the different serotypes. We compiled a list of 105 known and putative virulence genes of A. pleuropneumoniae from published literature and performed a diversity study of these genes at the nucleotide level, comparing serotypes 2 and 6. Sixty-two conserved virulence genes (Ͼ99% identity or fewer than three mismatches) were found between these two serotypes, while 28 virulence genes showed a larger degree of dissimilarity (Ͻ95% identity or more than 20 mismatches), including candidates like the apxIVA gene encoding RTX toxin and the cysI gene encoding NADPH-sulfite reductase hemoprotein. Capsular polysaccharides (CPS) produced by A. pleuropneumoniae are considered to be important virulence f...

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“…or to confirm, that control pigs stayed A.pp .-negative until the end of the study [33]. Bacterial species diagnostic was confirmed by PCR for the ApxIIA gene and by urease activity [34,35]. …”

Section: Methodsmentioning

confidence: 99%

Actinobacillus pleuropneumoniae challenge in swine: diagnostic of lung alterations by infrared thermography

Menzel

Beyerbach

Siewert³

et al. 2014

BMC Vet Res

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BackgroundActinobacillus pleuropneumoniae (A.pp.) is the causative agent of porcine pleuropneumonia leading to high economic losses in the pig industry. Infrared thermography (IRT) of the thorax might offer a new method to select swine with lung alterations for further diagnostics.In this study 50 german landrace pigs were infected with A.pp. in an established model for respiratory tract disease, while 10 healthy pigs served as control animals. To avoid drift errors during IR measurements absolute skin temperatures and temperature differences between a thoracal and an abdominal region were assessed for its diagnostic validity.ResultsIRT findings during the course of experimental A.pp.-infection were verified by computed tomography (CT) before and on days 4 and 21 after infection. Significant correlations were found between clinical scores, CT score and lung lesion score. Ambient temperature, body temperature and abdominal surface temperature were factors influencing the skin surface temperature of the thorax. On day 4 but not on day 21 after infection the right thoracal temperature was significantly higher and the difference between a thoracal region in the height of the left 10th vertebra and an abdominal region was significantly lower in infected pigs than in control pigs. At a cut off of 28°C of right thoracal temperature the specificity of the method was 100% (CI 95%: 69-100%) and the sensitivity 66% (CI 95%: 51-79%).At a cut off of 2°C temperature difference between thoracal and abdominal region on the left body site the specificity of the method was 100% (CI 95%: 69-100%) and the sensitivity 32% (CI 95%: 19-47%) with all control pigs detected negative.Orientation for lung biopsy by IRT resulted in 100% specificity and sensitivity (CI 95%: 69-100%) of bacteriological examination of tissue samples during the acute stage of infection.ConclusionIRT might be a valuable tool for the detection of inflammatory lung alterations in pigs, especially during the acute stage of infection and if ambient temperatures are constant during individual measurements. External and internal factors interfere with this method, so that its application in the field might be restricted to a selection of pigs for further diagnostic with adequate specificity.

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Development of a DIVA subunit vaccine against Actinobacillus pleuropneumoniae infection

Cited by 33 publications

References 33 publications

Proteomic and immunoproteomic characterization of a DIVA subunit vaccine against Actinobacillus pleuropneumoniae

Proteomic and immunoproteomic characterization of a DIVA subunit vaccine against Actinobacillus pleuropneumoniae

Draft Genome Sequences of Actinobacillus pleuropneumoniae Serotypes 2 and 6

Actinobacillus pleuropneumoniae challenge in swine: diagnostic of lung alterations by infrared thermography

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