Pleuropneumoniae Infection Research Articles

The Effect of Inactivated Mycobacterium Paratuberculosis Vaccine on the Response to a Heterologous Bacterial Challenge in Pigs.

Background: Vaccines may have non-specific effects, affecting resistance to heterologous pathogens. Veterinary vaccines have seldom been investigated for their non-specific effects. However, recent observational studies suggest that an inactivated paratuberculosis vaccine reduced all-cause mortality in goats and cattle.Aim: We tested if vaccination with a killed mycobacterial vaccine may have heterologous effects in swine (Sus domesticus), specifically on the pathogenic and clinical effects of a heterologous challenge with Actinobacillus pleuropneumoniae in young pigs.Methods: Newborn piglets were randomized to vaccination s.c. with the inactivated paratuberculosis vaccine Gudair (Zoetis Inc.) (n = 17) or no vaccine (n = 16). At 4–5 weeks after vaccination, all piglets were challenged intra-nasally with a high (Gudair: n = 8; control: n = 8) or a low (Gudair: n = 9; control: n = 8) dose of the gram-negative bacterium A. pleuropneumoniae causing acute porcine pleuropneumonia. The effect and severity of pathogen challenge was evaluated by measuring acute phase proteins C-reactive protein, haptoglobin and Porcine α1-acid glycoprotein, and by gross pathology 1 day post challenge. Specific and non-specific in vitro cytokine responses to vaccination were evaluated in whole blood before bacterial challenge.Results: The vaccine was immunogenic in the pigs as evidenced by increased IFN-γ responses to purified protein derivative of Mycobacterium paratuberculosis. However, Gudair vaccine did not affect IL-6 responses. The gross pathology of the lungs as well as the acute phase protein responses after the high A. pleuropneumoniae dose challenge was slightly increased in the vaccinated animals compared with controls, whereas this was not seen in the animals receiving the low-dose bacterial challenge.Conclusion: The inactivated paratuberculosis vaccine exacerbated the pathological and inflammatory effects of an experimental A. pleuropneumoniae infection in young pigs.

Comparison of pharmacokinetics of tilmicosin in healthy pigs and pigs experimentally infected with Actinobacillus pleuropneumoniae

ABSTRACTAim: To compare the pharmacokinetic profiles of tilmicosin, administered orally at a single dose of 20 mg/kg bodyweight, in healthy pigs and in pigs experimentally infected with Actinobacillus pleuropneumoniae.Methods: Twelve healthy crossbred pigs, aged approximately 8 weeks, were randomly assigned to uninfected and infected groups, with six pigs per group. Pigs in the infected group were inoculated intranasally with a bacterial suspension of A. pleuropneumoniae containing approximately 108 cfu. Each pig received a single oral dose of 20 mg/kg bodyweight of tilmicosin, given 3–4 hours after inoculation in infected pigs. Blood samples were collected before drug administration and up to 48 hours after tilmicosin administration. Concentrations of tilmicosin in plasma samples were determined by HPLC. Throughout the experimental period pigs were observed for signs of inappetence and clinical abnormalities. After sampling was complete pigs were subject to euthanasia and samples collected for gross and histopathology as well as microbiology.Results: Infected pigs showed signs of bradykinesia, nasal discharge dyspnoea, and coughing 1 hours after inoculation and A. pleuropneumoniae was cultured from the lungs of all infected pigs postmortem. Comparing pharmacokinetic parameters in uninfected and infected pigs, the maximum plasma concentration of tilmicosin was higher in uninfected pigs (1.17 (SD 0.17) vs. 0.96 (SD 0.17) µg/mL), the time to reach maximum concentration was shorter (1.53 (SD 0.23) vs. 2.40 (SD 0.37) hours), and the half-life of the absorption phase and half-life of the elimination phase were both shorter (0.66 (SD 0.08) vs. 1.00 (SD 0.27) hours) and (12.93 (SD 0.96) vs. 16.53 (SD 0.55) hours), respectively. The apparent volume of distribution was smaller in uninfected than infected pigs (1.91 (SD 0.22) vs. 2.16 (SD 0.21) L/kg). The relative bioavailability of tilmicosin in infected relative to uninfected pigs was 108.6 (SD 9.71)%.Conclusions and clinical relevance: The results of this study indicate that A. pleuropneumoniae infection significantly changed certain pharmacokinetic parameters of tilmicosin in pigs. In infected pigs tilmicosin exhibited a longer drug persistence and a better extent of absorption. These results indicate that it is necessary to monitor and adjust the dose of tilmicosin administration during the presence of pleuropneumonia. It is expected that this can optimise clinical efficacy and help avoid the development of resistance.

A bivalent fusion vaccine composed of recombinant Apx proteins shows strong protection against Actinobacillus pleuroneumoniae serovar 1 and 2 in a mouse model.

Actinobacillus pleuropneumonia (APP) causes porcine pleuropneumoniae, resulting in severe economic losses in the swine industry. Since there are diverse serotypes of APP, it is necessary for vaccines to induce cross-protection. In this report, we developed a bivalent fusion vaccine, the L vaccine composed of ApxIA and ApxIIA fragments. According to the experimental results of the L vaccine, recombinant protein specific-IgG antibody level increased significantly as well as Apx toxin specific-IgG antibody, suggesting toxin-neutralizing effect. Also, the production of both IgG1 and IgG2a indicates this fusion vaccine induces Th1 and Th2 immune reactions. In addition, lymphocytes were proliferated and immune related-cytokines of TNF-α, IL-12, IFN-γ and IL-5 were detected in the serum after the vaccination. The L vaccine showed a perfect cross-protection against APP serovar 1 and 2 that each secrete different Apx exotoxins. These findings reveal that the fusion L vaccine induces specific humoral and cellular immunity, leading to a perfect cross-protection against A. pleuropneumoniae infections in a murine model.

Antimicrobial resistance, biofilm formation and virulence reveal Actinobacillus pleuropneumoniae strains' pathogenicity complexity

Porcine pleuropneumonia is an important cause of lowered productivity and economic loss in the pig industry worldwide, associated primarily with Actinobacillus pleuropneumoniae infection. Its colonization and persistence within the upper respiratory tract of affected pigs depends upon interactions between a number of genetically controlled virulence factors, such as pore-forming repeats-in-toxin exoproteins, biofilm formation, and antimicrobial resistance. This study investigated correlations between biofilm-forming capacity, antimicrobial resistance, and virulence of A. pleuropneumoniae obtained from clinical outbreaks of disease, using a Galleria mellonella alternative infection model. Results suggest that virulence is diverse amongst the 21 strains of A. pleuropneumoniae examined and biofilm formation correlated with genetic control of antimicrobial resistance.

Respiratory explants as a model to investigate early events of contagious bovine pleuropneumonia infection

Contagious bovine pleuropneumonia (CBPP) is a severe disease caused by Mycoplasma mycoides subsp. mycoides (Mmm). Knowledge on CBPP pathogenesis is fragmented and hampered by the limited availability of laboratory animal and in vitro models of investigation. The purpose of the present study is to assess respiratory explants as useful tools to study the early stages of CBPP. Explants were obtained from trachea, bronchi and lungs of slaughtered cattle, tested negative for Mycoplasma spp. and for the major bacterial and viral respiratory pathogens. The interaction of Mmm with explant cells was studied by immunohistochemistry (IHC), double-labelling indirect immunofluorescence (DLIIF) and laser scanning confocal microscopy (LSCM). Mmm capability to survive and proliferate within the explants was evaluated by standard microbiological procedures. Finally, the putative cellular internalization of Mmm was further investigated by the gentamicin invasion assay. IHC and DLIIF indicated that Mmm can colonize explants, showing a marked tropism for lower airways. Specifically, Mmm was detected on/inside the bronchiolar and alveolar epithelial cells, the alveolar macrophages and the endothelial cells. The interaction between Mmm and explant cells was abolished by the pre-incubation of the pathogen with bovine anti-Mmm immune sera. Mmm was able to survive and proliferate in all tracheal, bronchial and lung explants, during the entire time course of the experiments. LSCM and gentamicin invasion assay both confirmed that Mmm can enter non-phagocytic host cells. Taken together, our data supports bovine respiratory explants as a promising tool to investigate CBPP, alternative to cattle experimental infection.

In vivo testing of novel vaccine prototypes against Actinobacillus pleuropneumoniae

Actinobacillus pleuropneumoniae (A. pleuropneumoniae) is a Gram-negative bacterium that represents the main cause of porcine pleuropneumonia in pigs, causing significant economic losses to the livestock industry worldwide. A. pleuropneumoniae, as the majority of Gram-negative bacteria, excrete vesicles from its outer membrane (OM), accordingly defined as outer membrane vesicles (OMVs). Thanks to their antigenic similarity to the OM, OMVs have emerged as a promising tool in vaccinology. In this study we describe the in vivo testing of several vaccine prototypes for the prevention of infection by all known A. pleuropneumoniae serotypes. Previously identified vaccine candidates, the recombinant proteins ApfA and VacJ, administered individually or in various combinations with the OMVs, were employed as vaccination strategies. Our data show that the addition of the OMVs in the vaccine formulations significantly increased the specific IgG titer against both ApfA and VacJ in the immunized animals, confirming the previously postulated potential of the OMVs as adjuvant. Unfortunately, the antibody response raised did not translate into an effective protection against A. pleuropneumoniae infection, as none of the immunized groups following challenge showed a significantly lower degree of lesions than the controls. Interestingly, quite the opposite was true, as the animals with the highest IgG titers were also the ones bearing the most extensive lesions in their lungs. These results shed new light on A. pleuropneumoniae pathogenicity, suggesting that antibody-mediated cytotoxicity from the host immune response may play a central role in the development of the lesions typically associated with A. pleuropneumoniae infections.

Identification and characterization of serovar-independent immunogens in Actinobacillus pleuropneumoniae

Despite numerous actions to prevent disease, Actinobacillus pleuropneumoniae (A. pleuropneumoniae) remains a major cause of porcine pleuropneumonia, resulting in economic losses to the swine industry worldwide. In this paper, we describe the utilization of a reverse vaccinology approach for the selection and in vitro testing of serovar-independent A. pleuropneumoniae immunogens. Potential immunogens were identified in the complete genomes of three A. pleuropneumoniae strains belonging to different serovars using the following parameters: predicted outer-membrane subcellular localization; ≤ 1 trans-membrane helices; presence of a signal peptide in the protein sequence; presence in all known A. pleuropneumoniae genomes; homology with other well characterized factors with relevant data regarding immunogenicity/protective potential. Using this approach, we selected the proteins ApfA and VacJ to be expressed and further characterized, both in silico and in vitro. Additionally, we analysed outer membrane vesicles (OMVs) of A. pleuropneumoniae MIDG2331 as potential immunogens, and compared deletions in degS and nlpI for increasing yields of OMVs compared to the parental strain. Our results indicated that ApfA and VacJ are highly conserved proteins, naturally expressed during infection by all A. pleuropneumoniae serovars tested. Furthermore, OMVs, ApfA and VacJ were shown to possess a high immunogenic potential in vitro. These findings favour the immunogen selection protocol used, and suggest that OMVs, along with ApfA and VacJ, could represent effective immunogens for the prevention of A. pleuropneumoniae infections in a serovar-independent manner. This hypothesis is nonetheless predictive in nature, and in vivo testing in a relevant animal model will be necessary to verify its validity.

Pig Lung Immune Cytokine Response to the Swine Influenza Virus and the Actinobacillus Pleuropneumoniae Infection.

IntroductionThe aim of this study was to evaluate and compare the local innate immune response to the swine influenza virus (SIV) and Actinobacillus pleuropneumoniae (App) infection in pigs.Material and MethodsThe study was performed on 37 seven-week-old pigs, divided into four groups: App-infected (n=11), App+SIV-infected (n=11), SIV-infected (n=11), and control (n=4). Lung samples were collected, following euthanasia, on the 2nd and 4th dpi (three piglets per inoculated group) and on the 10th dpi (remaining inoculated and control pigs). Lung concentrations of IL-1β, IL-6, IL-8, TNF-α, IL-10, IFN-α, and IFN-γ were analysed with the use of commercial porcine cytokine ELISA kits.ResultsLung concentrations of IL-1β, IL-6, IL-8, TNF-α, IFN-α, and IFN-γ were induced in SIV-infected and App+SIV-infected pigs. In the lung tissue of App-infected pigs, only concentrations of IL-1β, IL-6, IL-8, and IFN-γ were elevated. Additionally, in App+SIV-infected pigs, significantly greater concentrations of IL-1β, IL-8, and IFN-α were found when compared with pigs infected with either SIV or App alone. In each tested group, the lung concentration of IL-10 remained unchanged during the entire study.ConclusionThe results of the study indicate that the experimental infection of pigs with SIV or App alone and co-infection with both pathogens induced a local lung inflammatory response. However, the local cytokine response was considerably higher in co-infected pigs compared to single-infected pigs.

English

Contagious bovine pleuropneumonia (CBPP) is a highly infectious cattle disease, which is widespread in pastoral areas of Africa and it is a major problem for Ethiopian livestock. A cross-sectional study on sero-epidemiology and risk factors for contagious bovine pleuropneumonia infection of cattle in Dello Mena and Sawena districts of Bale Zone was conducted from February, 2016 to May, 2016. Competitive Enzyme Linked Immunosorbent Assay test was used to analyze 384 cattle sera for contagious bovine pleuropneumonia antibodies and the overall sero-prevalence rate was determined to be 6.51%. A higher sero-prevalence rate of 8.26% was observed in Dello Mena district when compared to Sawena (3.89%) district. The prevalence in each kebele ranged from 0 to 15.91% and the highest prevalence (15.91%) was found at Hi-Oda, located in Dello Mena district. A Chi-square computed statistical analysis indicated that age (χ2=5.738; p<0.017), sex (χ2=11.105; p<0.001), breed (χ2=13.29; p<0.014), body condition score (χ2=6.063; p<0.048) and purpose of rearing cattle (χ2=14.877; p<0.001) were the major putative risk factors associated with the occurrence of contagious bovine pleuropneumonia in the study districts. The difference, however, was not statistically significant (p>0.05) for origin (Dello Mena/Sawena), contact with other herds (yes/no) and herd size (small/medium/large). Multivariable logistic regression statistical analysis revealed that age (adult/young), body condition score (good/medium/poor) and purpose of rearing cattle (beef/draft power/milk) were significantly associated with contagious bovine pleuropneumonia sero-positivity (p<0.05). Consequently, cattle rearing purpose (for beef/draft power/milk) was statistically identified as the major risk factor for contagious bovine pleuropneumonia  occurrence in relation to other factors and cattle(oxen/bull) used for the purpose of draft power were more likely to be infected by CBPP(Adjusted Odds Ratio=49.052). This study showed that the overall sero-epidemiology of contagious bovine pleuropneumonia in Bale Zone of Oromia Regional State was relatively high. This warrants the implementation of appropriate preventive and control measures to minimize the economic losses arising from the disease.   Key words: Cattle, contagious bovine pleuropneumonia, Dello Mena, risk factors, Sawena, seroepidemiology

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.

Thymol kills bacteria, reduces biofilm formation, and protects mice against a fatal infection of Actinobacillus pleuropneumoniae strain L20

Actinobacillus pleuropneumoniae is the causative agent of the highly contagious and deadly respiratory infection porcine pleuropneumonia, resulting in serious losses to the pig industry worldwide. Alternative to antibiotics are urgently needed due to the serious increase in antimicrobial resistance. Thymol is a monoterpene phenol and efficiently kills a variety of bacteria. This study found that thymol has strong bactericidal effects on the A. pleuropneumoniae 5b serotype strain, an epidemic strain in China. Sterilization occurred rapidly, and the minimum inhibitory concentration (MIC) is 31.25μg/mL; the A. pleuropneumoniae density was reduced 1000 times within 10min following treatment with 1 MIC. Transmission electron microscopy (TEM) analysis revealed that thymol could rapidly disrupt the cell walls and cell membranes of A. pleuropneumoniae, causing leakage of cell contents and cell death. In addition, treatment with thymol at 0.5 MIC significantly reduced the biofilm formation of A. pleuropneumoniae. Quantitative RT-PCR results indicated that thymol treatment significantly increased the expression of the virulence genes purC, tbpB1 and clpP and down-regulated ApxI, ApxII and Apa1 expression in A. pleuropneumoniae. Therapeutic analysis of a murine model showed that thymol (20mg/kg) protected mice from a lethal dose of A. pleuropneumoniae, attenuated lung pathological lesions. This study is the first to report the use of thymol to treat A. pleuropneumoniae infection, establishing a foundation for the development of new antimicrobials.

Sequestrum in a Lymph Node in Contagious Bovine Pleuropneumonia (CBPP) Infection without Lung Lesions: A Case Report

Contagious bovine pleuropneumonia (CBPP) is a highly contagious disease of ruminants caused by Mycoplasma mycoides subsp. mycoides (Mmm), with a major impact on livestock production and a potential for rapid spread. It is manifested by anorexia, fever and respiratory signs such as dyspnoea, polypnoea, cough and nasal discharges in cattle. Lesions of CBPP are generally confined to the organs of the thoracic cavity except in young calves where inflammation of the limb joints (usually the carpal and tarsal joints), sometimes seen with increased fluid. The characteristic type of lesions seen in the organs generally include; pleuritis, hypertrophy of the lymph nodes, unilateral marbling of the lungs in acute disease and sequestra formation in subacute to chronic cases. This case study report describes the presence of a well-developed sequestrum in a peri-bronchial lymph node at the trachea bifurcation in an animal artificially infected with Mmm causing CBPP. The epidemiological role such lesions play in the transmission of the disease in the field is not known. This requires further investigation to ascertain whethersuch lesions are implicated in the transmission and perpetuation of disease. In addition such lesions would be missed at post-mortem in an outbreak as they are small and in unexpected organs leading to misdiagnosis of the disease.

Lactobacillus coryniformis Causing Pulmonary Infection in a Patient with Metastatic Small Cell Carcinoma: Case Report and Review of Literature on Lactobacillus Pleuro-Pulmonary Infections

Lactobacilli are normal commensals of the gastrointestinal and female genital tract. Due to its low virulence these bacteria are known to cause opportunistic infections. They cause mostly bacteraemia with or without endocarditis and rarely cause pleuro-pulmonary infection. We report a case of Lactobacillus coryniformis pleuro-pulmonary infection and review 14 previously reported cases of lactobacilli causing pleuro-pulmonary infections. Our patient had small cell carcinoma with metastasis. All the 14 cases had pre-existing risk factors like immunosuppresion, cancer or chronic disease. There was no consensus on choice of antimicrobial agents to be used. Different species of lactobacilli were involved. Available susceptibility data showed that lactobacilli species were more susceptible to ampicillin, erythromycin, gentamycin, and clindamycin and decreased to ceftriaxone, ciprofloxacin and trimethoprim-sulphamethoxazole. Isolation of Lactobacillus species from a case of pleuro-pneumonia infection could be a marker of poor long-term prognosis. The diagnosis of these infections requires both microbiologist and clinical correlation to rule out contamination.

Apa2H1, the first head domain of Apa2 trimeric autotransporter adhesin, activates mouse bone marrow-derived dendritic cells and immunization with Apa2H1 protects against Actinobacillus pleuropneumoniae infection.

Actinobacillus pleuropneumoniae is the causative pathogen of porcine pleuropneumonia, which results in large economic losses in the pig industry worldwide. There are, however, no effective subunit vaccines are available in the market owing to the various serotypes and the absence of cross-protection against this pathogen. Therefore, the selection of protective components is of great significance for vaccine development. We previously showed that trimeric autotransporter adhesins are important virulence factors of A. pleuropneumoniae. To determine the potential role in vaccine development of the functional head domain (Apa2H1) of Apa2, a trimeric autotransporter adhesin found in A. pleuropneumoniae, we obtained nature-like trimeric Apa2H1 using a prokaryotic expression system and co-culture of Apa2H1 with bone marrow derived dendritic cells (BMDCs) in vitro resulted in maturation of BMDCs, characterised by the up-regulation of CD83, MHC-II, CCR7, ICAM-I and the increased expression of factors related to B lymphoid cells stimulation, such as proliferation-inducing ligand (APRIL), B lymphocyte stimulator (BLyS) and B cell activating factor (BAFF). The in vivo results showed that vaccination with Apa2H1 resulted in the robust production of antigen-specific antibodies, modestly induced mixed Th1 and Th2 immunity, impaired bacterial colonization and dissemination, and improved mouse survival rates. This study is the first to show that Apa2H1 is antigenic and can be used as a component of a subunit vaccine against A. pleuropneumoniae infection, providing valuable reference material for the development of an effective vaccine against A. pleuropneumoniae.

Attenuated Actinobacillus pleuropneumoniae double-deletion mutant S-8∆clpP/apxIIC confers protection against homologous or heterologous strain challenge.

Background Actinobacillus pleuropneumoniae is the etiological agent of porcine pleuropneumonia, which leads to large economic losses to the swine industry worldwide. In this study, S-8△clpP△apxIIC, a double-deletion mutant of A. pleuropneumoniae was constructed, and its safety and protective efficacy were evaluated in pigs.ResultsThe S-8△clpP△apxIIC mutant exhibited attenuated virulence in a murine (BALB/c) model, and caused no detrimental effects on pigs even at a dose of up to 1.0 × 109 CFU. Furthermore, the S-8△clpP△apxIIC mutant was able to induce a strong immune response in pigs, which included high levels of IgG1 and IgG2, stimulated gamma interferon (IFN-γ), interleukin 12 (IL-12), and interleukin 4 (IL-4) production, and conferred effective protection against the lethal challenge with A. pleuropneumoniae serovars 7 or 5a. The pigs in the S-8△clpP△apxIIC immunized groups have no lesions and reduced bacterial loads in the lung tissue after challenge.ConclusionsThe data obtained in this study suggest that the S-8△clpP△apxIIC mutant can serve as a highly immunogenic and potential live attenuated vaccine candidate against A. pleuropneumoniae infection.

Immunoprotective Efficacy of Six In vivo-Induced Antigens against Actinobacillus pleuropneumoniae as Potential Vaccine Candidates in Murine Model.

Six in vivo-induced (IVI) antigens—RnhB, GalU, GalT, Apl_1061, Apl_1166, and HflX were selected for a vaccine trial in a mouse model. The results showed that the IgG levels in each immune group was significantly higher than that of the negative control (P < 0.001). Except rRnhB group, proliferation of splenocytes was observed in all immunized groups and a relatively higher proliferation activity was observed in rGalU and rGalT groups (P < 0.05). In the rGalT vaccinated group, the proportion of CD4+ T cells in spleen was significant higher than that of negative control (P < 0.05). Moreover, proportions of CD4+ T cells in other vaccinated groups were all up-regulated to varying degrees. Up-regulation of both Th1 (IFN-γ, IL-2) and Th2 (IL-4) cytokines were detected. A survival rate of 87.5, 62.5, and 62.5% were obtained among rGalT, rAPL_1166, and rHflX group, respectively while the remaining three groups was only 25%. Histopathological analyses of lungs indicated that surviving animals from the vaccinated groups showed relatively normal pulmonary structure alveoli. These findings confirm that IVI antigens used as vaccine candidates provide partial protection against Actinobacillus pleuropneumoniae infection in a mouse model, which could be used as potential vaccine candidates in piglets.

Economic value of mitigating Actinobacillus pleuropneumoniae infections in pig fattening herds

Actinobacillus pleuropneumoniae infections (APP) can cause severe economic losses in pig production. Swine producers can prevent the disease from occurring and spreading. However, to be profitable for the producer, the anticipated benefits of a mitigation policy should exceed its cost.In this study, a dynamic programming model and multivariate normal approximation of general stochastic epidemic model (SIR) was developed, and policies for managing fattening pigs under the risk of APP infection were evaluated. The following disease mitigation options were considered: 1) cleaning including washing, drying and disinfection procedures (for efficient and inefficient cleaning), 2) cleaning and vaccination (with high, medium and low efficacy of vaccine), and 3) cleaning and medication (antibiotics use). The results were calculated for five different levels of APP prevalence, where the disease can occur at ten levels of severity.The economic value of mitigation policy depended on the prevalence and severity of disease as well as on the efficiency of the implemented mitigation protocol. The results suggest that inefficient cleaning caused substantial economic losses which can reach up to €20.2 per pig space unit per year.The cleaning and vaccination policy was economically superior over the policy of only cleaning when the prevalence of disease was high (>0.3), there was a severe clinical disease and the medium or high efficacy vaccine was available. Moreover, even when vaccination costs were decreased the vaccination policy was not economically favored in situations with low disease prevalence among herds (0.1 and 0.3).In cases of severe disease, the cleaning and medication policy resulted in higher economic benefits than cleaning or cleaning and vaccination with low and medium efficacy vaccine. With a high prevalence (>0.3) and substantial decrease in the average daily gain (ADG), the joint application of cleaning and medication was economically less favorable than cleaning combined with the use of the high efficacy vaccine. Therefore, the availability of high efficacy vaccine could benefit farms facing the severe forms of APP and help to reduce the use of antibiotics.To avoid substantial economic losses, pig producers should not ignore cleaning procedures, which due to low efficacy of available vaccines and concerns regarding excessive use of antibiotics and antibiotic resistance in animal production should be the primary policy to mitigate APP.

Identification and characterization of a novel stress-responsive outer membrane protein Lip40 from Actinobacillus pleuropneumoniae

BackgroundActinobacillus pleuropneumoniae, a Gram-negative bacterium, is the causative agent of porcine pleuropneumonia, a highly contagious and often fatal disease. Because current vaccines confer limited protection against A. pleuropneumoniae infection, the development of more effective vaccines is urgently required. The identification of immunogenic and protective antigens, such as an outer-membrane lipoprotein, will advance this purpose.ResultsSixty putative lipoproteins were predicted from the genomic sequence of A. pleuropneumoniae using multiple algorithms. Here, we focused on the characteristics of the putative lipoprotein Lip40 from A. pleuropneumoniae strain SLW01 (serovar 1). Lip40 shares sequence similarity with many bacterial lipoproteins, and the structural prediction of Lip40 suggests that it is similar to A. pleuropneumoniae TbpB. The N-terminus of Lip40 contains an interesting tandemly repeated sequence, Q(E/D/P)QPK. Real-time RT–PCR indicated that the expression of lip40 was significantly upregulated at 42 °C, at 16 °C, and under anaerobic conditions. Recombinant Lip40 (rLip40) produced in Escherichia coli BL21(DE3) was specifically recognized by porcine convalescent serum directed against A. pleuropneumoniae. Lip40 was confirmed to localize at the bacterial outer membrane, and its expression was significantly stimulated when A. pleuropneumoniae was cultured under various stress conditions. Lip40 also protected 75 % of mice from fatal virulent A. pleuropneumoniae infection.ConclusionsThe immunogenic outer-membrane protein Lip40 is stress responsive, protects mice against infection, and might be a virulence determinant. Further investigation of Lip40 should expedite vaccine development and provide insight into the pathogenesis of A. pleuropneumoniae.Electronic supplementary materialThe online version of this article (doi:10.1186/s12896-015-0199-8) contains supplementary material, which is available to authorized users.

Identification of Actinobacillus pleuropneumoniae Genes Preferentially Expressed During Infection Using In Vivo-Induced Antigen Technology (IVIAT).

Porcine pleuropneumonia is an infectious disease caused by Actinobacillus pleuropneumoniae. The identification of A. pleuropneumoniae genes, specially expressed in vivo, is a useful tool to reveal the mechanism of infection. IVIAT was used in this work to identify antigens expressed in vivo during A. pleuropneumoniae infection, using sera from individuals with chronic porcine pleuropneumonia. Sequencing of DNA inserts from positive clones showed 11 open reading frames with high homology to A. pleuropneumoniae genes. Based on sequence analysis, proteins encoded by these genes were involved in metabolism, replication, transcription regulation, and signal transduction. Moreover, three function-unknown proteins were also indentified in this work. Expression analysis using quantitative real-time PCR showed that most of the genes tested were up-regulated in vivo relative to their expression levels in vitro. IVI (in vivoinduced) genes that were amplified by PCR in different A. pleuropneumoniae strains showed that these genes could be detected in almost all of the strains. It is demonstrated that the identified IVI antigen may have important roles in the infection of A. pleuropneumoniae.

Immunomodulatory effects of tulathromycin on apoptosis, efferocytosis, and proinflammatory leukotriene B4 production in leukocytes from Actinobacillus pleuropneumoniae-or zymosan-challenged pigs.

To investigate the anti-inflammatory and immunomodulatory properties of tulathromycin in vitro and in experimental models of Actinobacillus pleuropneumoniae-induced pleuropneumonia and zymosan-induced pulmonary inflammation in pigs. Blood samples from six 8- to 30-week-old healthy male pigs for the in vitro experiment and sixty-five 3-week-old specific pathogen-free pigs. Neutrophils and monocyte-derived macrophages were isolated from blood samples. Isolated cells were exposed to tulathromycin (0.02 to 2.0 mg/mL) for various durations and assessed for markers of apoptosis and efferocytosis. For in vivo experiments, pigs were inoculated intratracheally with A pleuropneumoniae, zymosan, or PBS solution (control group) with or without tulathromycin pretreatment (2.5 mg/kg, IM). Bronchoalveolar lavage fluid was collected 3 and 24 hours after inoculation and analyzed for proinflammatory mediators, leukocyte apoptosis, and efferocytosis. In vitro, tulathromycin induced time- and concentration-dependent apoptosis in neutrophils, which enhanced their subsequent clearance by macrophages. In the lungs of both A pleuropneumoniae- and zymosan-challenged pigs, tulathromycin promoted leukocyte apoptosis and efferocytosis and inhibited proinflammatory leukotriene B4 production, with a concurrent reduction in leukocyte necrosis relative to that of control pigs. Tulathromycin also attenuated the degree of lung damage and lesion progression in A pleuropneumoniae-inoculated pigs. Tulathromycin had immunomodulatory effects in leukocytes in vitro and anti-inflammatory effects in pigs in experimental models of A pleuropneumoniae infection and nonmicrobial-induced pulmonary inflammation. These data suggested that in addition to its antimicrobial properties, tulathromycin may dampen severe proinflammatory responses and drive resolution of inflammation in pigs with microbial pulmonary infections.