Actinobacillus Pleuropneumoniae Serotype Research Articles

Histopathological lesions of Actinobacillus pleuropneumoniae serotype 8 in infected pigs.

This study aimed to report, for the first time, histopathological lesions caused by an outbreak of acute Actinobacillus pleuropneumoniae serotype 8 infections in two farms in Cyprus. Lung tissue samples were collected from two different affected farms (a total of eight samples) for bacterial culture, multiplex polymerase chain reaction (PCR)-based serotyping and histopathological evaluation. Severe respiratory clinical signs, vomiting, anorexia, sudden deaths, a morbidity rate of around 25.00% and a mortality rate of over 60.00% in the fattening stage were reported. Macroscopic lesions included acute to subacute fibrotic, hemorrhagic and necrotizing pneumonia with occasionally encapsulated nodule-like abscesses and fibrous pleuritis. Histopathological evaluation revealed fibrous exudate between alveolar spaces and connective tissue, areas of necrosis mixed with alveolar macrophages, lymphocytes, plasma cells and necrotic leukocytes surrounding colonies of cocci. The bronchial and bronchiolar epithelia were degenerated and replaced by eosinophilic cell debris mixed with inflammatory cells. Several arteries and capillaries were clotted and/or infiltrated by inflammatory cells. In conclusion, these A. pleuropneumoniae serotype 8 cases were accompanied by acute illness, death and more pronounced bronchitis and bronchiolitis.

Revealing Genomic Insights of the Unexplored Porcine Pathogen Actinobacillus pleuropneumoniae Using Whole Genome Sequencing.

ABSTRACTActinobacillus pleuropneumoniae (APP) is the causative agent of pleuropneumonia in pigs, one of the most relevant bacterial respiratory diseases in the swine industry. To date, 19 serotypes have been described based on capsular polysaccharide typing with significant virulence dissimilarities. In this study, 16 APP isolates from Spanish origin were selected to perform antimicrobial susceptibility tests and comparative genomic analysis using whole genome sequencing (WGS). To obtain a more comprehensive worldwide molecular epidemiologic analyses, all APP whole genome assemblies available at the National Center for Biotechnology Information (NCBI) at the time of the study were also included. An in-house in silico PCR approach enabled the correct serotyping of unserotyped or incorrectly serotyped isolates and allowed for the discrimination between serotypes 9 and 11. A pangenome analysis identified the presence or absence of gene clusters to be serotype specific, as well as virulence profile analyses targeting the apx operons. Antimicrobial resistance genes were correlated to the presence of specific plasmids. Altogether, this study provides new insights into the genetic variability within APP serotypes, correlates phenotypic tests with bioinformatic analyses and manifests the benefits of populated databases for a better assessment of diversity and variability of relatively unknown pathogens. Overall, genomic comparative analysis enhances the understanding of transmission and epidemiological patterns of this species and suggests vertical transmission of the pathogen, including the resistance genes, within the Spanish integrated systems.IMPORTANCE Pleuropneumonia is one of the most relevant respiratory infections in the swine industry. Despite Actinobacillus pleuropneumoniae (APP) being one of the most important pathogens in the pig production, this is the first comparative study including all available whole genome sequencing data from NCBI. Moreover, this study also includes 16 APP isolates of Spanish origin with known epidemiological relationships through vertical integrated systems. Genomic comparisons provided a deeper understanding of molecular and epidemiological knowledge between different APP serotypes. Furthermore, determination of resistance and toxin profiles allowed correlation with the presence of mobile genetic elements and specific serotype, respectively.

Draft Genome Sequences of Two Clinical Actinobacillus pleuropneumoniae Serotype 19 Strains from Pigs in Switzerland.

ABSTRACTActinobacillus pleuropneumoniae serotype 19 is a very recently described new serotype with a novel type II capsule synthesis locus. Here, we report the draft genome sequences of two Actinobacillus pleuropneumoniae serotype 19 strains with a serogroup 3/6/8/12/15 O-antigen locus that were isolated in 2018 and 2021 from two different pig farms in Switzerland.

Serologic monitoring of herds with and without bacterin vaccination for Actinobacillus pleuropneumoniae

Objective: Investigate diagnostic serology for Actinobacillus pleuropneumoniae (APP) infections in naturally infected and vaccinated pigs. Materials and methods: The APP status of 12 farms (A-L) was established by lung cultures and isolate serotyping. Screening enzyme-linked immunosorbent assay (ELISA) detected antibodies to ApxIV antigen or multiple APP serotypes. Serotype-specific ELISAs were conducted for serotypes 5 and 7. Seven groups of farm F pigs (serotype 7) were moved to farm K (serotype 5). Autogenous vaccines (V1/V2) prepared from APP serotype 5 cultures from farm K and a commercial, killed APP vaccine (V3) containing serotypes 1, 7, and 15 were used to vaccinate pigs in each group twice or thrice at 3-week intervals, commencing at 10 weeks of age. Blood samples were analyzed with ELISAs specific for serotype 5 and ApxI and ApxII toxins. Serum titers were compared using an analysis of variance. Results: Serotypes 5, 7, 12, or 15 were present in lung cultures. The ApxIV screening ELISA and mix-serotype ELISA regularly detected serotypes 5, 7, and 15. Serotype 12 infections were detected in the mix-serotype ELISA, but not in the ApxIV assays. The serotype 5 or 7 specific ELISA regularly detected herd infections with the relevant serotype. Serotype 5 titers of pigs vaccinated with V1/V2 thrice were higher than those dosed twice with the equivalent volume (P < .05). Pigs receiving V3 showed no serotype 5 antibody response. The ApxI and II titers in V1/V2-vaccinated pigs were higher than controls. Implications: Screening and serotype-specific ELISAs verified APP status. Repeated serotype-specific autogenous APP vaccine doses provided a strong antibody response.

Exposure to benzylpenicillin after different dosage regimens in growing pigs

BackgroundPenicillin is important for treatment of pigs, but data on its absorption and disposition in pigs are sparse. This is reflected by the variation in recommended dosages in the literature. Inadequate dosage may lead to treatment failure and selection of resistant bacteria. To optimize treatment regimens, plasma exposure to benzylpenicillin for two sustained release formulations of procaine benzylpenicillin for intramuscular administration was studied in growing pigs by means of tandem mass spectrometry (UPLC–MS/MS). One formulation was an aqueous suspension, Ethacilin® vet (ETH), and the other an oily suspension, Ultrapen vet (UPA). Benzylpenicillin exposure after intravenous administration of potassium benzylpenicillin was also explored. Exposure profiles were first studied after single administrations of the approved dosages in healthy pigs and then after repeated administration of different dosages in pigs inoculated intranasally with an Actinobacillus pleuropneumoniae serotype 2 strain.ResultsAfter intravenous administration of benzylpenicillin (n = 6), maximum plasma concentration (Cmax), 1860–9318 µg/L, was observed after 15 min. At four h, plasma concentrations decreased to 15–76 µg/L. After intramuscular administration of ETH (n = 6) Cmax, 1000–4270 µg/L, was observed within one h (tmax) in 5 pigs but at four h in one pig. Cmax for UPA (n = 6), 910–3220 µg/L, was observed within one h in three pigs, but at four or 24 h in three pigs. For both ETH and UPA, the terminal phase was characterized by slow decline compared with intravenous administration. Repeated administration of different dosages of ETH and UPA in pigs inoculated with A. pleuropneumoniae (n = 54) showed that the approved dose for UPA (30 mg/kg, qd) but not for ETH (20 mg/kg, qd) gave adequate plasma exposure for bacteria with a penicillin MIC of 500 µg/L. However, more frequent dosing of ETH (bid) or increased dosage gave an adequate exposure.ConclusionsThe approved dosage of ETH provided insufficient plasma exposure for adequate therapy of infections caused by A. pleuropneumoniae or other bacteria with a penicillin MIC of 500 µg/L. More frequent ETH dosing (bid) or an increased dosage would improve exposure. The approved dosage of UPA however provided adequate exposure.

Enhancement of Apx Toxin Production in Actinobacillus pleuropneumoniae Serotypes 1, 2, and 5 by Optimizing Culture Condition.

Actinobacillus pleuropneumoniae (APP) is a causative agent of porcine pleuropneumonia. Therefore, the development of an effective vaccine for APP is necessary. Here, we optimized the culture medium and conditions to enhance the production yields of Apx toxins in APP serotype 1, 2, and 5 cultures. The use of Mycoplasma Broth Base (PPLO) medium improved both the quantity and quality of the harvested Apx toxins compared with Columbia Broth medium. Calcium chloride (CaCl2) was first demonstrated as a stimulation factor for the production of Apx toxins in APP serotype 2 cultures. Cultivation of APP serotype 2 in PPLO medium supplemented with 10 μg/ml of nicotinamide adenine dinucleotide (NAD) and 20 mM CaCl2 yielded the highest levels of Apx toxins. These findings suggest that the optimization of the culture medium and conditions increases the concentration of Apx toxins in the supernatants of APP serotype 1, 2, and 5 cultures and may be applied for the development of vaccines against APP infection.

Construction and immunization with double mutant ΔapxIBD Δpnp forms of Actinobacillus pleuropneumoniae serotypes 1 and 5.

Actinobacillus pleuropneumoniae (APP) causes a form of porcine pleuropneumonia that leads to significant economic losses in the swine industry worldwide. The apxIBD gene is responsible for the secretion of the ApxI and ApxII toxins and the pnp gene is responsible for the adaptation of bacteria to cold temperature and a virulence factor. The apxIBD and pnp genes were deleted successfully from APP serotype 1 and 5 by transconjugation and sucrose counter-selection. The APP1ΔapxIBDΔpnp and APP5ΔapxIBDΔpnp mutants lost hemolytic activity and could not secrete ApxI and ApxII toxins outside the bacteria because both mutants lost the ApxI- and ApxII-secreting proteins by deletion of the apxIBD gene. Besides, the growth of these mutants was defective at low temperatures resulting from the deletion of pnp. The APP1ΔapxIBDΔpnp and APP5ΔapxIBDΔpnp mutants were significantly attenuated compared with wild-type ones. However, mice vaccinated intraperitoneally with APP5ΔapxIBDΔpnp did not provide any protection when challenged with a 10-times 50% lethal dose of virulent homologous (APP5) and heterologous (APP1) bacterial strains, while mice vaccinated with APP1ΔapxIBDΔpnp offered 75% protection against a homologous challenge. The ΔapxIBDΔpnp mutants were significantly attenuated and gave different protection rate against homologous virulent wild-type APP challenging.

Effect Found in the Combination of Kitasamycin with Other Antibiotics Used against <i>Actinobacillus pleuropneumoniae</i>

Background: Combinations of antibiotics are used to increase the therapeutic options via the simultaneous activities of two compounds. The goals of combination therapy are to resolve the bacterial infection while reducing the treatment cost. The use of kitasamycin is used frequently and in addition to other antibiotics, the desired combinations were not found. Methods: Fields strains: we use serotypes 1, 3, 5 y 7. Microbial culturing: The serotypes 1, 3, 5 y 7 was identified. Antibiotics and Reagents: The antibiotics used were kitasamycin; enrofloxacin and oxolinic acid. The following combinations were tested: a) kitasamycin with enrofloxacin; b) kitasamycin with norfloxacin; and c) kitasamycin with oxolinic acid. Preparation of bacterial suspensions: The cultures were adjusted to a concentration of 5 colony-forming units per ml (CFU/ml) using the same sterile culture medium as the diluent. Determination of the fractional inhibitory concentrations (FICs): To test each combination and their interactions and to calculate the fractional inhibitory concentration (FIC) of each antibiotic. Calculation of the index of the fractional minimum inhibitory concentration (FMIC): The FMIC index was calculated by applying the following formula: FIC index = FIC of antibiotic A/FIC of antibiotic B. Results: What was found showed that: A combination of kitasamycin and norfloxacin has been shown to have an indifferent effect. A combination of kitasamycin and enrofloxacin had antagonistic effects against all of the Actinobacillus pleuropneumoniae serotypes studied. Finally, a combination of kitasamycin and oxolinic acid had antagonist effects against serotypes 1, 3 and 5 and a synergistic effect against serotype 7. Conclusion: The “in vitro” combination Kitasamycin-Norfloxacin showed an indifferent effect, and combination Kitasamycin and Enrofloxacin demonstrated an antagonistic interaction between these two antibiotics. A combination of Kitasamycin and Oxolinic Acid had synergistic effect against serotype 7.

375 Identification of QTL associated with antibody response to common infectious diseases in commercial sows

Abstract The objective of this study was to perform genome-wide association studies (GWAS) to identify Quantitative Trait Loci (QTL) associated with antibody response to infectious diseases in commercial sows. A total of 2,848 Large White x Landrace replacement gilts were sourced from 17 high-health multipliers (7 breeding companies; BC) and introduced to 23 commercial farms with a history of common pig diseases, following the standard acclimation procedures with an average of 53 animals per entry group (CG). Serum was used to quantify antibody response to swine influenza virus (SIV), Mycoplasma hyopneumoniae (MH), porcine circovirus type-2 (PCV2), and 8 serotypes of Actinobacillus pleuropneumoniae (APP1-3, 5, 7, 10, 12, and 13) at entry (S/PEntry), following acclimation (S/PAcclimation), and during parities 1 (S/PParity1) and 2 (S/PParity2). All animals were genotyped for 38,191 SNPs. GWAS was performed using BayesB (pi=0.99), with the fixed effect of CG and the random effects of SNPs in the model. For APP, QTL were only identified at S/PAcclimation; on SSC14 (2Mb) for APP3, APP7, APP10, and APP13 that explained 5.6, 4.7, 2.8, and 3.6% of the genetic variance, respectively. A gene within this QTL region is SYK, involved in the control of immune-receptors. For APP5, a QTL that explained 4.2% of the genetic variance was identified on SSC4 (105Mb), which co-localizes with two genes associated with immune-response: SIKE1and NRAS. For SIV, no QTL was identified. A QTL on SSC7 (130-131Mb) was identified for MH (S/PParity1, 5.1%) and PCV2 (S/PEntry, 34%; S/PAcclimation, 43.4%). These results provide new information on the genetic basis of response to infectious diseases in sows. The identified QTL have the potential to be used to select for improved immune response. The authors thanks PigGen Canada, Genome Canada, and the Canadian Swine Health Board for financial support, and the late Dr. Stephen Bishop for his scientific contributions.

PSIII-5 Accuracy of genomic prediction of antibody response to common infectious diseases in commercial sows

Abstract Previous results indicated that antibody response to PRRSV has moderate genomic prediction accuracy; however, little is known about this for other common infectious diseases. Therefore, the objective of this study was to estimate the accuracy of genomic prediction for antibody response to infectious diseases in commercial sows. A total of 2,848 Large White x Landrace replacement gilts were sourced from 17 high-health multipliers (7 breeding companies; BC) and introduced to 23 commercial farms with a history of common diseases, following standard acclimation procedures. Serum was used to quantify antibody response to swine influenza virus (SIV), Mycoplasma hyopneumoniae (MH), porcine circovirus type 2 (PCV2), and 8 serotypes of Actinobacilluspleuropneumoniae(APP1-3, 5, 7, 10, 12, and 13) at entry (S/PEntry), following acclimation (S/PAcclimation), and during parities 1 (S/PParity1) and 2 (S/PParity2). All animals were genotyped for 38,191 SNPs. Genomic prediction was performed using BayesB (pi=0.99), with the fixed effect of CG and random effects of SNPs included in the model. Training and validation were performed using 7-fold cross-validation, with data from each BC used as the validation dataset in one-fold. In general, prediction accuracies were low: SIV, from 0.13 (S/PAcclimation) to 0.26 (S/PParity1); MH, -0.07 (S/PAcclimation) to 0.13 (S/PParity2); PCV2, 0.04 (S/PParity1) to 0.32 (S/PAcclimation); APP, -0.08 (S/PEntry, APP10) to 0.26 (S/PAcclimation, APP7). At each point, average accuracies were 0.06 for S/PEntry, 0.09 for S/PAcclimationand S/PParity1, and 0.08 for S/PParity2, showing small increases in accuracy after the acclimation period. Among diseases, average accuracies ranged from 0.01 (APP1) to 0.22 (PCV2). Results show that, overall, the accuracy of genomic prediction of antibody response to common infectious diseases in commercial gilts is limited. The authors thank PigGen Canada, Genome Canada, and the Canadian Swine Health Board for financial support, and the late Dr. Stephen Bishop for his scientific contributions.

An anti-Propionibacterium acnes antibody shows heterologous resistance to an Actinobacillus pleuropneumoniae infection independent of neutrophils in mice

Porcine contagious pleuropneumonia is a highly fatal respiratory disease that is caused by Actinobacillus pleuropneumoniae (APP) and results in tremendous economic losses for the pig breeding industry worldwide. Previous studies have demonstrated that Propionibacterium acnes (PA) could effectively prevent APP infection in mice and pigs. The humoral immune response played a primary role during this process and anti-PA antibody could mediate macrophages to kill the bacteria. However, the role of neutrophils in this process is currently unknown. In this study, mice were injected with cyclophosphamide to deplete neutrophils and then passively immunized with anti-PA serum or negative serum. Mice were subsequently challenged with APP serotype 1. The results showed that the mice exhibited less bacterial colonization, less lung damage, and a high survival rate, which were immunized with the anti-PA antibody whether neutrophils were depleted or not. Worse still, the presence of neutrophils increased the damage to the mice after challenge. These results suggest that the activity of the anti-PA antibody against APP infection was independent of neutrophils. These findings have important significance for understanding the mechanisms of humoral immunity conferred by heterologous immunization and lay a good foundation for preventing APP infection.

B cell cross-epitope of Propionibacterium acnes and Actinobacillus pleuropneumonia selected by phage display library can efficiently protect from Actinobacillus pleuropneumonia infection

Contagious porcine pleuropneumonia (CPP), caused by Actinobacillus pleuropneumoniae (APP), is a highly transmissible and fatal respiratory illness that causes tremendous economic losses for the pig breeding industry worldwide. Propionibacterium acnes (PA) has a strong cross-reaction with anti-APP1 and anti-APP5 serum and can efficiently prevent APP infection, which was fortuitously found in researching the differential gene between the different APP serotypes. There seems to be some natural cross-protection between PA and APP. To identify the common epitope, the phage display library of a PA whole genome was constructed, whose size is 105. The DNA sequence of the positive clone was determined after three rounds of biopanning, and ten common protein types were identified and the epitope was predicted by computer software. Six peptide epitopes were selected and synthesized for further analysis. Among these epitopes, Ba1, Bb5 and C1 could bind to anti-PA serum and anti-APP1 serum and vice versa. Furthermore, the IgG and IL-4 levels and CD4+/CD8+ T cell ratios in the Ba1, Bb5 and C1 groups were significantly higher than that in the control group, indicating that the epitopes could trigger an immune response, which was mainly humoral immunity. Moreover, Ba1 and Bb5 equally protected 80% of mice from a fatal dose of APP1 infection compared with the control group. Mice could resist APP1 and APP5 challenge after being treated with the combination of Ba1 and Bb5, with survival rates of 80% and 90%, respectively. These findings suggest that the PA epitope confers antigenicity and can heterologously resist to the APP infection. This finding provides a novel strategy for preventing APP infection.

Efficacy Evaluation of Combination Vaccine of Recombinant C-Terminal Fragments of ApxIA, ApxIIA and ApxIIIA in Piglets

The efficacy of the combination vaccine of the individual C-terminal fragments of ApxIA, ApxIIA and ApxIIIA of Actinobacillus pleuropneumoniae (APP) was evaluated in piglets. Twenty piglets were divided equally into 2 groups (n=10). All piglets were intramuscularly primed at 4 week-of-age (0 week post prime inoculation (WPPI)) and were intramuscularly boosted at 6 week-of-age (2 WPPI). Group A piglets were inoculated with sterile PBS and group B piglets were inoculated with the combination vaccine. Concentrations of each of the C-terminal fragment-specific IgG as determined by ELISA were significantly higher in group B than in group A from 2 WPPI until the end of this study. Clinical signs were observed from only 10% of group B piglets after the challenge with the mixture of APP serotypes 1, 2 and 5 at 4 WPPI, while 50% of group A piglets were protected against APP infections. Overall, intramuscular inoculation with the vaccine candidate can efficiently protect piglets against APP infection.

Frequency of Th17 cells correlates with the presence of lung lesions in pigs chronically infected with Actinobacillus pleuropneumoniae

Porcine contagious pleuropneumonia caused by Actinobacillus pleuropneumoniae (APP) remains one of the major causes of poor growth performance and respiratory disease in pig herds. While the role of antibodies against APP has been intensely studied, the porcine T cell response remains poorly characterized. To address this, pigs were intranasally infected with APP serotype 2 and euthanized during the acute phase [6–10 days post-infection (dpi)] or the chronic phase of APP infection (27–31 dpi). Lymphocytes isolated from blood, tonsils, lung tissue and tracheobronchial lymph nodes were analyzed by intracellular cytokine staining (ICS) for IL-17A, IL-10 and TNF-α production after in vitro stimulation with crude capsular extract (CCE) of the APP inoculation strain. This was combined with cell surface staining for the expression of CD4, CD8α and TCR-γδ. Clinical records, microbiological investigations and pathological findings confirmed the induction of a subclinical APP infection. ICS-assays revealed the presence of APP-CCE specific CD4+CD8αdim IL-17A-producing T cells in blood and lung tissue in most infected animals during the acute and chronic phase of infection and a minor fraction of these cells co-produced TNF-α. APP-CCE specific IL-17A-producing γδ T cells could not be found and APP-CCE specific IL-10-producing CD4+ T cells were present in various organs but only in a few infected animals. The frequency of identified putative Th17 cells (CD4+CD8αdimIL-17A+) in lung and blood correlated positively with lung lesion scores and APP-specific antibody titers during the chronic phase. These results suggest a potential role of Th17 cells in the immune pathogenesis of APP infection.

Molecular serotyping and antimicrobial resistance profiles of Actinobacillus pleuropneumoniae isolated from pigs in South Korea

ABSTRACTBackground: Actinobacillus pleuropneumoniae (APP) causes porcine pleuropneumonia (PP).Objective: Serotypes and antimicrobial resistance patterns in APP isolates from pigs in Korea were examined.Methods: Sixty-five APP isolates were genetically serotyped using standard and multiplex PCR (polymerase chain reaction). Antimicrobial susceptibilities were tested using the standardized disk-agar method. PCR was used to detect β-lactam, gentamicin and tetracycline-resistance genes. The random amplified polymorphic DNA (RAPD) patterns were determined by PCR.Results: Korean pigs predominantly carried APP serotypes 1 and 5. Among 65 isolates, one isolate was sensitive to all 12 antimicrobials tested in this study. Sixty-two isolates was resistant to tetracycline and 53 isolates carried one or five genes including tet(B), tet(A), tet(H), tet(M)/tet(O), tet(C), tet(G) and/or tet(L)-1 markers. Among 64 strains, 9% and 26.6% were resistance to 10 and three or more antimicrobials, respectively. Thirteen different antimicrobial resistance patterns were observed and RAPD analysis revealed a separation of the isolates into two clusters: cluster II (6 strains resistant to 10 antimicrobials) and cluster I (the other 59 strains).Conclusion: Results show that APP serotypes 1 and 5 are the most common in Korea, and multi-drug resistant strains are prevalent. RAPD analysis demonstrated that six isolates resistant to 10 antimicrobials belonged to the same cluster.

Draft Genome Sequences of Six Actinobacillus pleuropneumoniae Serotype 8 Brazilian Clinical Isolates: Insight into New Applications.

Actinobacillus pleuropneumoniae is the causative agent of swine pleuropneumonia, a highly contagious disease associated with pigs of all ages that results in severe economic losses to the industry. Here, we report for the first time six genome sequences of A. pleuropneumoniae clinical isolates of serotype 8, found worldwide.

Actinobacillus pleuropneumoniae seroprevalence in farmed wild boars in Finland.

The aim of this study was to estimate the prevalence of serum antibodies to Actinobacillus pleuropneumoniae (APP) in Finnish farmed wild boars and to evaluate basic risk factors for seropositivity of the agent. A sampling frame was compiled based on the national record of wild boar farmers (n = 117), and 26 herds were included in the study. Serum samples (n = 206) were obtained from all animals slaughtered from these herds during the 2007-2008 period. Indirect enzyme-linked immunosorbent assay techniques were used to detect the possible presence of antibodies toward 12 serotypes of APP. Altogether, 1,442 analyses were performed, out of which 35 (2.4%) exceeded the positive cut-off value. Correspondingly, 26 of the pigs (12.6%) were found to be seropositive for any serotype of APP. The apparent animal prevalence of the cross-reacting APP serotypes 1, 9, and 11 was 3.9% (95% confidence interval [CI] 1-7%); it was 0.5% (CI 0-1%) for serotype 2; 3.4% (CI 1-6%) for serotypes 3, 6, and 8; 7.3% (CI 4-11%) for serotypes 4 and 7; 0.5% (CI 0-1%) for serotype 10; and 1.5% (CI 0-3%) for serotype 12. No antibodies against serotype 5s were detected. The apparent farm prevalence of APP serotypes 1,9, and 11 was 15.4% (CI 2-29%); that of serotype 2 was 3.8% (CI 0-11%); that of serotypes 3, 6, and 8 was 11.5% (CI 0-24%); that of serotypes 4 and 7 was 34.6% (CI 16-53%); that of serotype 10 was 3.8% (CI 0-11%); and that of serotype 12 was 7.7% (CI 0-18%). In conclusion, the prevalence of serum antibodies to different APP serotypes was low in farmed wild boar. Risk factor analysis did not reveal any significant animal- or herd-level risk factors for seropositivity of the agent.

Characterization of the omlA gene from different serotypes of Actinobacillus pleuropneumoniae: a new insight into an old approach

The OmlA protein is a virulence factor of Actinobacillus pleuropneumoniae, an important pathogen in pigs. The polymorphisms present in the omlA gene sequence of 15 reference serotypes of A. pleuropneumoniae and non-serotypable isolates were assessed to determine the possible evolutionary relationship among them and to validate the importance of this gene as a molecular marker for the characterization of this bacterium. Divergence among the 15 serotypes of A. pleuropneumoniae probably resulted initially from two major evolutionary events that led to subsequent differentiation into nine groups. This differentiation makes it possible to characterize most of the serotypes by using bionformatics, thereby avoiding problems with immunological cross-reactivity. A conserved α-helix common to all the serotypes was most likely involved in connecting the protein to the outer membrane and acting as a signal peptide. A previously unknown gene duplication was also identified and could contribute to the genetic variability that makes it difficult to serotype some isolates. Our data support the importance of the omlA gene in the biology of A. pleuropneumoniae and provide a new area of research into the OmlA protein.

Experimental Identification of Actinobacillus pleuropneumoniae Strains L20 and JL03 Heptosyltransferases, Evidence for a New Heptosyltransferase Signature Sequence

We experimentally identified the activities of six predicted heptosyltransferases in Actinobacillus pleuropneumoniae genome serotype 5b strain L20 and serotype 3 strain JL03. The initial identification was based on a bioinformatic analysis of the amino acid similarity between these putative heptosyltrasferases with others of known function from enteric bacteria and Aeromonas. The putative functions of all the Actinobacillus pleuropneumoniae heptosyltrasferases were determined by using surrogate LPS acceptor molecules from well-defined A. hydrophyla AH-3 and A. salmonicida A450 mutants. Our results show that heptosyltransferases APL_0981 and APJL_1001 are responsible for the transfer of the terminal outer core D-glycero-D-manno-heptose (D,D-Hep) residue although they are not currently included in the CAZY glycosyltransferase 9 family. The WahF heptosyltransferase group signature sequence [S(T/S)(GA)XXH] differs from the heptosyltransferases consensus signature sequence [D(TS)(GA)XXH], because of the substitution of D261 for S261, being unique.

Genetic variability in swine leukocyte antigen class II and Toll-like receptors affects immune responses to vaccination for bacterial infections in pigs

The genes encoding swine leukocyte antigen (SLA) and Toll-like receptor (TLR) are highly polymorphic in pig populations, and likely have influences on infection and the effects of vaccination. We explored the associations of different genotypes of SLA class II and of the genes TLR1, TLR4, TLR5, and TLR6 with antibody responses after vaccination against Erysipelothrix rhusiopathiae (ER) and Actinobacillus pleuropneumoniae (APP) serotypes 1, 2, and 5 in 191 Duroc pigs maintained under specific pathogen-free conditions. We demonstrated close relationships between SLA class II and ER antibody response and between TLR genes other than TLR4 and APP antibody responses. Pigs with specific haplotypes in SLA class II or TLR5 showed decreased antibody response to ER vaccination or increased responses to APP2 and APP5 vaccination, respectively. It might be possible to breed for responsiveness to vaccination and to implement new vaccine development strategies unaffected by genetic backgrounds of pigs.