Evaluation the kill rate and mutant selection window of danofloxacin against Actinobacillus pleuropneumoniae in a peristaltic pump model.

BACKGROUND: Actinobacillus pleuropneumoniae is a serious pathogen in pigs. The abundant application of antibiotics has resulted in the gradual emergence of drugresistant bacteria, which has seriously affected treatment of disease. To aid measures to prevent the emergence and spread of drug-resistant bacteria, herein, the kill rate and mutant selection window (MSW) of danofloxacin (DAN) against A. pleuropneumoniae were evaluated. METHODS: For the kill rate study, the minimum inhibitory concentration (MIC) was tested using the micro dilution broth method and time-killing curves of DAN against A. pleuropneumoniae grown in tryptic soy broth (TSB) at a series drug concentrations (from 0 to 64 MIC) were constructed. The relationships between the kill rate and drug concentrations were analyzed using a Sigmoid Emax model during different time periods. For the MSW study, the MIC99 (the lowest concentration that inhibited the growth of the bacteria by ≥ 99%) and mutant prevention concentration (MPC) of DAN against A. pleuropneumoniae were measured using the agar plate method. Then, a peristaltic pump infection model was established to simulate the dynamic changes of DAN concentrations in pig lungs. The changes in number and sensitivity of A. pleuropneumoniae were measured. The relationships between pharmacokinetic/pharmacodynamic parameters and the antibacterial effect were analyzed using the Sigmoid Emax model. RESULTS: In kill rate study, the MIC of DAN against A. pleuropneumoniae was 0.016 µg/mL. According to the kill rate, DAN exhibited concentration-dependent antibacterial activity against A. pleuropneumoniae. A bactericidal effect was observed when the DAN concentration reached 4-8 MIC. The kill rate increased constantly with the increase in DAN concentration, with a maximum value of 3.23 Log10 colony forming units (CFU)/mL/h during the 0-1 h period. When the drug concentration was in the middle part of the MSW, drugresistant bacteria might be induced. Therefore, the dosage should be avoided to produce a mean value of AUC24h/MIC99 (between 31.29 and 62.59 h. The values of AUC24h/MIC99 to achieve bacteriostatic, bactericidal, and eradication effects were 9.46, 25.14, and > 62.59 h, respectively. CONCLUSION: These kill rate and MSW results will provide valuable guidance for the use of DAN to treat A. pleuropneumoniae infections.

Naringin attenuates Actinobacillus pleuropneumoniae-induced acute lung injury via MAPK/NF-κB and Keap1/Nrf2/HO-1 pathway.

Actinobacillus pleuropneumoniae (APP) causes porcine pleuropneumonia (PCP), which is clinically characterized by acute hemorrhagic, necrotizing pneumonia, and chronic fibrinous pneumonia. Although many measures have been taken to prevent the disease, prevention and control of the disease are becoming increasingly difficult due to the abundance of APP sera, weak vaccine cross-protection, and increasing antibiotic resistance in APP. Therefore, there is an urgent need to develop novel drugs against APP infection to prevent the spread of APP. Naringin (NAR) has been reported to have an excellent therapeutic effect on pulmonary diseases, but its therapeutic effect on lung injury caused by APP is not apparent. Our research has shown that NAR was able to alleviate APP-induced weight loss and quantity of food taken and reduce the number of WBCs and NEs in peripheral blood in mice; pathological tissue sections showed that NAR was able to prevent and control APP-induced pathological lung injury effectively; based on the establishment of an in vivo/in vitro model of APP inflammation, it was found that NAR was able to play an anti-inflammatory role through inhibiting the MAPK/NF-κB signaling pathway and exerting anti-inflammatory effects; additionally, NAR activating the Nrf2 signalling pathway, increasing the secretion of antioxidant enzymes Nqo1, CAT, and SOD1, inhibiting the secretion of oxidative damage factors NOS2 and COX2, and enhancing the antioxidant stress ability, thus playing an antioxidant role. In summary, NAR can relieve severe lung injury caused by APP by reducing excessive inflammatory response and improving antioxidant capacity.

Tea Polyphenols Protects Tracheal Epithelial Tight Junctions in Lung during Actinobacillus pleuropneumoniae Infection via Suppressing TLR-4/MAPK/PKC-MLCK Signaling.

Actinobacillus pleuropneumoniae (APP) is the causative pathogen of porcine pleuropneumonia, a highly contagious respiratory disease in the pig industry. The increasingly severe antimicrobial resistance in APP urgently requires novel antibacterial alternatives for the treatment of APP infection. In this study, we investigated the effect of tea polyphenols (TP) against APP. MIC and MBC of TP showed significant inhibitory effects on bacteria growth and caused cellular damage to APP. Furthermore, TP decreased adherent activity of APP to the newborn pig tracheal epithelial cells (NPTr) and the destruction of the tight adherence junction proteins ß-catenin and occludin. Moreover, TP improved the survival rate of APP infected mice but also attenuated the release of the inflammation-related cytokines IL-6, IL-8, and TNF-α. TP inhibited activation of the TLR/MAPK/PKC-MLCK signaling for down-regulated TLR-2, TLR4, p-JNK, p-p38, p-PKC-α, and MLCK in cells triggered by APP. Collectively, our data suggest that TP represents a promising therapeutic agent in the treatment of APP infection.

Optimal regimens based on PK/PD cutoff evaluation of ceftiofur against Actinobacillus pleuropneumoniae in swine.

BACKGROUND: Actinobacillus pleuropneumoniae formerly known as Haemophilus pleuropneumoniae, can cause pleuropneumoniae in pigs, which lead to significant mortality. Ceftiofur was the first cephalosporin antibiotic used in animals, which was effective against gram-negative and gram-positive bacterium. This study aimed to formulate a rational dosage strategy and review the preceding recommended dosage based on PK/PD modeling and Establish Clinical breakpoint of ceftiofur against Actinobacillus pleuropneumoniae based on the pharmacodynamic-pharmacokinetic cutoff. RESULTS: The epidemiologic cutoff value was 0.125 µg/mL. The results of the pharmacodynamic study showed that the MICs of BW39 were 0.5 µg/mL and 1 µg/mL in vitro and ex-vivo, respectively. The minimal bactericidal concentrations (MBCs) under in vitro and ex vivo conditions were both 1 µg/mL. The time-killing profiles of ceftiofur against BW39 were time-dependent with a partly concentration-dependent pattern. Based on the inhibitory sigmoid Emax model, the AUC24 h/MIC values for the bacteriostatic, bactericidal, and elimination effects in serum were 45.73, 63.83, and 69.04 h for healthy pigs separately. According to the Monte Carlo simulation, the COPD was calculated as 2 µg/mL, and the optimized dosage regimen of ceftiofur against Actinobacillus pleuropneumoniae to achieve bacteriostatic, bactericidal, and elimination effects over 24 h was 2.13, 2.97, and 3.42 mg/kg for the 50% target attainment rate (TAR) and 2.47, 3.21, and 3.70 mg/kg for the 90% TAR respectively. CONCLUSIONS: In conclusion, we reveal the EOFF and PK/PD cutoff values of ceftiofur against A. pleuropneumoniae in piglets. However, with the paucity of clinical data for ceftiofur to establish a clinical cutoff against A. pleuropneumoniae, the PK/PD cutoff value of 2 µg/mL will be recommended as surrogate. According to the PK/PD data and the MIC distribution in China, the single bactericidal dose was 3.21 mg/kg for the 90% target, which would be more able to cure Actinobacillus pleuropneumoniae and avoid the emergence of resistance for clinical ceftiofur use in piglet.

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

Aim: 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.

Impact of growth matrix on pharmacodynamics of antimicrobial drugs for pig pneumonia pathogens.

BACKGROUND: The most widely used measure of potency of antimicrobial drugs is Minimum Inhibitory Concentration (MIC). MIC is usually determined under standardised conditions in broths formulated to optimise bacterial growth on a species-by-species basis. This ensures comparability of data between laboratories. However, differences in values of MIC may arise between broths of differing chemical composition and for some drug classes major differences occur between broths and biological fluids such as serum and inflammatory exudate. Such differences must be taken into account, when breakpoint PK/PD indices are derived and used to predict dosages for clinical use. There is therefore interest in comparing MIC values in several broths and, in particular, in comparing broth values with those generated in serum. For the pig pneumonia pathogens, Actinobacillus pleuropneumoniae and Pasteurella multocida, MICs were determined for three drugs, florfenicol, oxytetracycline and marbofloxacin, in five broths [Mueller Hinton Broth (MHB), cation-adjusted Mueller Hinton Broth (CAMHB), Columbia Broth supplemented with NAD (CB), Brain Heart Infusion Broth (BHI) and Tryptic Soy Broth (TSB)] and in pig serum. RESULTS: For each drug, similar MIC values were obtained in all broths, with one exception, marbofloxacin having similar MICs for three broths and 4-5-fold higher MICs for two broths. In contrast, for both organisms, quantitative differences between broth and pig serum MICs were obtained after correction of MICs for drug binding to serum protein (fu serum MIC). Potency was greater (fu serum MIC lower) in serum than in broths for marbofloxacin and florfenicol for both organisms. For oxytetracycline fu serum:broth MIC ratios were 6.30:1 (P. multocida) and 0.35:1 (A. pleuropneumoniae), so that potency of this drug was reduced for the former species and increased for the latter species. The chemical composition of pig serum and broths was compared; major matrix differences in 14 constituents did not account for MIC differences. Bacterial growth rates were compared in broths and pig serum in the absence of drugs; it was concluded that broth/serum MIC differences might be due to differing growth rates in some but not all instances. CONCLUSIONS: For all organisms and all drugs investigated in this study, it is suggested that broth MICs should be adjusted by an appropriate scaling factor when used to determine pharmacokinetic/pharmacodynamic breakpoints for dosage prediction.

What is the true in vitro potency of oxytetracycline for the pig pneumonia pathogens Actinobacillus pleuropneumoniae and Pasteurella multocida?

The pharmacodynamics of oxytetracycline was determined for pig respiratory tract pathogens, Actinobacillus pleuropneumoniae and Pasteurella multocida. Indices of potency were determined for the following: (i) two matrices, broth and pig serum; (ii) five overlapping sets of twofold dilutions; and (iii) a high strength starting culture. For A. pleuropneumoniae, minimum inhibitory concentration (MIC) was similar for the two matrices, but for P. multocida, differences were marked and significantly different. MIC and minimum bactericidal concentration (MBC) serum: broth ratios for A. pleuropneumoniae were 0.83:1 and 1.22:1, respectively, and corresponding values for P. multocida were 22.0:1 and 7.34:1. For mutant prevention concentration (MPC) serum: broth ratios were 0.79:1 (A. pleuropneumoniae) and 20.9:1 (P. multocida). These ratios were corrected for serum protein binding to yield fraction unbound (fu) serum: broth MIC ratios of 0.24:1 (A. pleuropneumoniae) and 6.30:1 (P. multocida). Corresponding fu serum: broth ratios for MPC were almost identical, 0.23:1 and 6.08:1. These corrections for protein binding did not account for potency differences between serum and broth for either species; based on fu serum MICs, potency in serum was approximately fourfold greater than predicted for A. pleuropneumoniae and sixfold smaller than predicted for P. multocida. For both broth and serum and both bacterial species, MICs were also dependent on initial inoculum strength. The killing action of oxytetracycline had the characteristics of codependency for both A. pleuropneumoniae and P. multocida in both growth media. The in vitro potency of oxytetracycline in pig serum is likely to be closer to the in vivo plasma/serum concentration required for efficacy than potency estimated in broths.

The influence of Actinobacillus pleuropneumoniae infection on tulathromycin pharmacokinetics and lung tissue disposition in pigs.

A tulathromycin concentration and pharmacokinetic parameters in plasma and lung tissue from healthy pigs and Actinobacillus pleuropneumoniae (App)-infected pigs were compared. Tulathromycin was administered intramuscularly (i.m.) to all pigs at a single dose of 2.5 mg/kg. Blood and lung tissue samples were collected during 33 days postdrug application. Tulathromycin concentration in plasma and lung was determined by high-performance liquid chromatography with tandem mass spectrometry (LC-MS/MS) method. The mean maximum plasma concentration (Cmax ) in healthy pigs was 586 ± 71 ng/mL, reached by 0.5 h, while the mean value for Cmax of tulathromycin in infected pigs was 386 ± 97 ng/mL after 0.5 h. The mean maximum tulathromycin concentration in lung of healthy group was calculated as 3412 ± 748 ng/g, detected at 12 h, while in pigs with App, the highest concentration in lung was 3337 ± 937 ng/g, determined at 48 h postdosing. The higher plasma and lung concentrations in pigs with no pulmonary inflammation were observed at the first time points sampling after tulathromycin administration, but slower elimination with elimination half-life t1/2el  = 126 h in plasma and t1/2el  = 165 h in lung, as well as longer drug persistent in infected pigs, was found.

Pharmacokinetic/pharmacodynamic evaluation of marbofloxacin in the treatment of Haemophilus parasuis and Actinobacillus pleuropneumoniae infections in nursery and fattener pigs using Monte Carlo simulations.

This study evaluated the theoretical clinical outcome of three marbofloxacin posology regimens in two groups of pigs (weaners and fatteners) for the treatment of Actinobacillus pleuropneumoniae (App) and Haemophilus parasuis (Hp) infection and the appearance of resistant bacteria due to the antibiotic treatment. The probability of target attainment (PTA) for pharmacokinetic/pharmacodynamics (PK/PD) ratios associated with clinical efficacy and with the appearance of antimicrobial resistance for fluoroquinolones at each minimum inhibitory concentration (MIC) or mutant prevention concentration (MPC) were calculated, respectively. The cumulative fraction of response (CFR) was calculated for the three posology regimens against App and they ranged from 91.12% to 96.37% in weaners and from 93% to 97.43% in fatteners, respectively. In the case of Hp, they ranged from 80.52% to 85.14% in weaners and from 82.01% to 88.49% in fatteners, respectively. Regarding the PTA of the PK/PD threshold associated with the appearance of antimicrobial resistance, results showed that marbofloxacin would prevent resistances in most of the animals up to the MPC value of 1 µg/mL.

A Florfenicol-Resistant Plasmid Shuttling Between Actinobacillus pleuropneumoniae and Glaesserella parasuis.

Porcine contagious pleuropneumonia, caused by Actinobacillus pleuropneumoniae, has resulted in significant economic losses to the swine industry. Although antibiotics are commonly employed to control this disease, their widespread use or misuse can lead to the development of antibiotic resistance in A. pleuropneumoniae. Consequently, it is crucial to conduct antimicrobial susceptibility testing on clinical isolates. In our study, we identified one strain of A. pleuropneumoniae with resistance to florfenicol and extracted a 5919 bp plasmid named pAPPJY, which confers florfenicol resistance. Sequence analysis revealed that the plasmid contains four open reading frames, namely rep, antioxin vbha family protein, floR, and a partial copy of lysr. Although a few variations in gene position were observed, the plasmid sequence exhibits a high degree of similarity to other florfenicol-resistant plasmids found in Glaesserella parasuis and A. pleuropneumoniae. Therefore, it is possible that the pAPPJY plasmid functions as a shuttle, facilitating the spread of florfenicol resistance between G. parasuis and A. pleuropneumoniae. In addition, partial recombination may occur during bacterial propagation. In conclusion, this study highlights the horizontal transmission of antibiotic resistance among different bacterial species through plasmids, underscoring the need for increased attention to antibiotic usage.

Rhein kills Actinobacillus pleuropneumoniae, reduces biofilm formation, and effectively treats bacterial lung infections in mice.

Background. Actinobacillus pleuropneumoniae, a member of the Pasteurellaceae family, is known for its highly infectious nature and is the primary causative agent of infectious pleuropneumonia in pigs. This disease poses a considerable threat to the global pig industry and leads to substantial economic losses due to reduced productivity, increased mortality rates, and the need for extensive veterinary care and treatment. Due to the emergence of multi-drug-resistant strains, Chinese herbal medicine is considered one of the best alternatives to antibiotics due to its unique mechanism of action and other properties. As a type of Chinese herbal medicine, Rhein has the advantages of a wide antibacterial spectrum and is less likely to develop drug resistance, which can perfectly solve the limitations of current antibacterial treatments.Methods. The killing effect of Rhein on A. pleuropneumoniae was detected by fluorescence quantification of differential expression changes of key genes, and scanning electron microscopy was used to observe the changes in A. pleuropneumoniae status after Rhein treatment. Establishing a mouse model to observe the treatment of Rhein after A. pleuropneumoniae infection.Results. Here, in this study, we found that Rhein had a good killing effect on A. pleuropneumoniae and that the MIC was 25 µg ml-1. After 3 h of action, Rhein (4×MIC) completely kills A. pleuropneumoniae and Rhein has good stability. In addition, the treatment with Rhein (1×MIC) significantly reduced the formation of bacterial biofilms. Therapeutic evaluation in a murine model showed that Rhein protects mice from A. pleuropneumoniae and relieves lung inflammation. Quantitative RT-PCR (Quantitative reverse transcription polymerase chain reaction is a molecular biology technique that combines both reverse transcription and polymerase chain reaction methods to quantitatively detect the amount of a specific RNA molecule) results showed that Rhein treatment significantly downregulated the expression of the IL-18 (Interleukin refers to a class of cytokines produced by white blood cells), TNF-α, p65 and p38 genes. Along with the downregulation of genes such as IL-18, it means that Rhein has an inhibitory effect on the expression of these genes, thereby reducing the activation of inflammatory cells and the production of inflammatory mediators. This helps reduce inflammation and protects tissue from further damage.Conclusions. This study reports the activity of Rhein against A. pleuropneumoniae and its mechanism, and reveals the ability of Rhein to treat A. pleuropneumoniae infection in mice, laying the foundation for the development of new drugs for bacterial infections.

Actinobacillus ureae may be a critical pathogen in patients with predispositions: A case report and review of the literature.

RATIONALE: Actinobacillus ureae (A. ureae) is an unusual commensal of human respiratory flora, rarely causing human infection. The predisposing factors, identification, clinical features, and antibiotic therapy of A. ureae are seldomly reported. Herein, we present a case of 64-year-old man affected by A. ureae pneumonia after intracranial surgery. PATIENT CONCERNS AND DIAGNOSES: A 64-year-old male was admitted with vomiting, drowsiness, and a severe disturbance of consciousness and was later diagnosed with cerebral hemorrhage by computed tomography images. After a craniocerebral surgery, the patient suffered from intractable pneumonia, experiencing treatment failure with multiple anti-bacterial agents. Sputum culture yield pure colonies of A. ureae, confirmed by matrix-assisted laser desorption/ionization time of flight and 16S rRNA gene sequencing. INTERVENTIONS: Minocycline (100 mg p.o. per 12 hours) with a course of 15 days was administrated for this patient. OUTCOMES: The respiratory symptoms, presenting as intermittent coughing with purulent and yellowish sputum, were gone. A 3-month follow-up examination showed a complete resolution of radiological findings. LESSONS: Clinically, the actual incidence of A. ureae pneumonia may be higher than that we generally recognized, and clinicians should consider A. ureae as a possible etiologic agent in patients with predispositions. Currently, A. ureae may be susceptible to penicillin, ampicillin, and third-generation cephalosporins. Other antibacterial agents, such as tetracycline, amoxicillin/clavulanic acid, and aminoglycosides also respond well and can be a choice in the treatment of A. ureae infections.

In vitro efficacy of diallyl sulfides against the periodontopathogen Aggregatibacter actinomycetemcomitans.

The in vitro antibacterial effects of diallyl sulfide (DAS) against the Gram-negative periodontopathogen Aggregatibacter actinomycetemcomitans, the key etiologic agent of the severe form of localized aggressive periodontitis and other nonoral infections, were studied. A. actinomycetemcomitans was treated with garlic extract, allicin, or DAS, and the anti-A. actinomycetemcomitans effects of the treatment were evaluated. Garlic extract, allicin, and DAS significantly inhibited the growth of A. actinomycetemcomitans (greater than 3 log; P < 0.01) compared to control cells. Heat inactivation of the garlic extracts significantly reduced the protein concentration; however, the antimicrobial effect was retained. Purified proteins from garlic extract did not exhibit antimicrobial activity. Allicin lost all its antimicrobial effect when it was subjected to heat treatment, whereas DAS demonstrated an antimicrobial effect similar to that of the garlic extract, suggesting that the antimicrobial activity of garlic extract is mainly due to DAS. An A. actinomycetemcomitans biofilm-killing assay performed with DAS showed a significant reduction in biofilm cell numbers, as evidenced by both confocal microscopy and culture. Scanning electron microscopy (SEM) analysis of DAS-treated A. actinomycetemcomitans biofilms showed alterations of colony architecture indicating severe stress. Flow cytometry analysis of OBA9 cells did not demonstrate apoptosis or cell cycle arrest at therapeutic concentrations of DAS (0.01 and 0.1 µg/ml). DAS-treated A. actinomycetemcomitans cells demonstrated complete inhibition of glutathione (GSH) S-transferase (GST) activity. However, OBA9 cells, when exposed to DAS at similar concentrations, showed no significant differences in GST activity, suggesting that DAS-induced GST inhibition might be involved in A. actinomycetemcomitans cell death. These findings demonstrate that DAS exhibits significant antibacterial activity against A. actinomycetemcomitans and that this property might be utilized for exploring its therapeutic potential in treatment of A. actinomycetemcomitans-associated oral and nonoral infections.

A.actinomycetemcomitans-induced periodontal disease promotes systemic and local responses in rat periodontium.

AIM: To characterize the histologic and cellular response to A. actinomycetemcomitans (Aa) infection. MATERIAL & METHODS: Wistar rats infected with Aa were evaluated for antibody response, oral Aa colonization, loss of attachment, PMN recruitment, TNF-α in the junctional epithelium and connective tissue, osteoclasts and adaptive immune response in local lymph nodes at baseline and 4, 5 or 6 weeks after infection. Some groups were given antibacterial treatment at 4 weeks. RESULTS: An antibody response against Aa occurred within 4 weeks of infection, and 78% of inoculated rats had detectable Aa in the oral cavity (p < 0.05). Aa infection significantly increased loss of attachment that was reversed by antibacterial treatment (p < 0.05). TNF-α expression in the junctional epithelium followed the same pattern. Aa stimulated high osteoclast formation and TNF-α expression in the connective tissue (p < 0.05). PMN recruitment significantly increased after Aa infection (p < 0.05). Aa also increased the number of CD8(+) T cells (p < 0.05), but not CD4(+) T cells or regulatory T cells (Tregs) (p > 0.05). CONCLUSION: Aa infection stimulated a local response that increased numbers of PMNs and TNF-α expression in the junctional epithelium and loss of attachment. Both TNF-α expression in JE and loss of attachment was reversed by antibiotic treatment. Aa infection also increased TNF-α in the connective tissue, osteoclast numbers and CD8(+) T cells in lymph nodes. The results link Aa infection with important characteristics of periodontal destruction.

Experimental arthritis exacerbates Aggregatibacter actinomycetemcomitans-induced periodontitis in mice.

AIM: This study aimed to investigate whether chronic antigen-induced arthritis (AIA) influences infection-induced periodontitis (PD) in mice and whether PD modifies the clinical course of AIA. The contribution of anti-TNF-α therapy was also evaluated. MATERIALS AND METHODS: The PD was induced in C57BL/6 mice by oral infection with Aggregatibacter actinomycetemcomitans. AIA was induced after infection. Anti-TNF-α and chlorhexidine therapies were used to investigate the role of TNF-α and oral infection on PD and AIA interaction. Maxillae, knee joints, lymph nodes and serum samples were used for histomorphometric, immunoenzymatic and/or real time-PCR analyses. RESULTS: Antigen-induced arthritis exacerbated alveolar bone loss triggered by PD infection. In contrast, PD did not influence AIA in the evaluated time-points. PD exacerbation was associated with enhanced production of IFN-γ in maxillae and expression of the Th1 transcription factor tBET in submandibular lymph nodes. Increased serum levels of IL-6 and C-reactive protein were also detected. Anti-TNF-α and antiseptic therapies prevented the development and exacerbation of infectious-PD. Anti-TNF-α therapy also resulted in reduced expression of IFN-γ, TNF-α and IL-17 in maxillae. CONCLUSIONS: Altogether, the current results indicate that the exacerbation of infection-induced PD by arthritis is associated with an alteration in lymphocyte polarization pattern and increased systemic immunoreactivity. This process was ameliorated by anti-TNF-α and antiseptic therapies.

Serotypes, antimicrobial susceptibility, and minimal inhibitory concentrations of Actionbacillus pleuropneumoniae isolated from slaughter pigs in Taiwan (2002-2007).

In total, 211 isolates of A. pleuropneumoniae were collected from pigs with hemorrhagic pneumonia at slaughterhouses during 2002-2007. Serotypes, antimicrobial susceptibility and minimum inhibitory concentration (MIC) values were determined for each isolate of A. pleuropneumoniae to 10 antimicrobial agents. Serovar 1 of A. pleuropneumoniae was predominant in Taiwan in 138 of the 211 isolates, followed by serovars 2 and 5. More than 90% of collected isolates were sensitive to ceftiofur, cephalothin, and chloramphenical. However, lincospectin and gentamicin were relatively less susceptible with sensitivities of only 2.4 and 5.7%, respectively. Additionally, ceftiofur had the highest in vitro activity with an MIC(50) of 2.2 µg/ml, followed by cephalothin (2.7 µg/ml) and chloramphenicol (7.9 µg/ml). Lincospectin had the least activity with MIC(50) and MIC(90) values of 73.9 and 114.5 µg/ml, respectively. The data indicate that ceftiofur and cephalothin were extremely active against A. pleuropneumoniae and with minimum MIC values. These drugs are suitable for controlling and treating hemorrhagic pleuropneumonia outbreaks in swine.

Gamithromycin in swine: Pharmacokinetics and clinical evaluation against swine respiratory disease.

The pharmacokinetics of gamithromycin were evaluated in 26 male castrated and female crossbred swine administered gamithromycin 15% w/v (Zactran®, Boehringer Ingelheim) intravenously at 6 mg/kg bodyweight or intramuscularly at 3, 6 or 12 mg/kg bodyweight. Blood samples were collected up to Day 10 to establish the plasma profile of gamithromycin, bioavailability and dose proportionality. When administered by intramuscular injection at 6 mg/kg BWT, pharmacokinetic parameters were as follows: area under the curve until last quantifiable plasma concentration, 5.13 ± 0.957 µg*hours/ml; maximum plasma concentration, 960 ± 153 ng/ml at 5 to 15 min; terminal half-life of 94.1 ± 20.4 hr. Absolute bioavailability was 92.2%. Increase in systemic exposure was proportional to the gamithromycin dose level over the range 3-12 mg/kg BWT. No gender-related statistically significant difference in exposure was observed. For clinical evaluation of Zactran® against swine respiratory disease, 305 pigs from six commercial farms in three countries in Europe with signs associated with Actinobacillus pleuropneumoniae and/or Haemophilus parasuis and/or Pasteurella multocida and/or Bordetella bronchiseptica were used. At each site, animals were treated once in a 1:1 ratio with a single intramuscular dose of Zactran® (6 mg gamithromycin/kg bodyweight) or Zuprevo® (4% w/v tildipirosin at 4 mg/kg bodyweight; MSD Animal Health) at the recommended dose respectively. Animals were observed and scored daily for 10 consecutive days for signs of swine respiratory disease (depression, respiration and rectal temperature), and animals presenting signs of clinical swine respiratory disease (Depression Score 3 and/or Respiratory Score 3 associated with Rectal Temperature > 40.0°C) were removed from the study and considered as treatment failure. Animals which remained in the study were individually assessed for 'treatment success' or 'treatment failure' (Depression Score ≥ 1 and Rectal Temperature > 40.0°C or Respiratory Score ≥ 1 and Rectal Temperature > 40.0°C). Using a non-inferiority hypothesis test (non-inferiority margin = 0.10), the proportion of treatment successes in the Zactran® group (97%) was equivalent to or better than that in the Zuprevo® group (93%).

Targeted delivery of a photosensitizer to Aggregatibacter actinomycetemcomitans biofilm.

The ability to selectively target specific biofilm species with antimicrobials would enable control over biofilm consortium composition, with medical applications in treatment of infections on mucosal surfaces that are colonized by a mixture of beneficial and pathogenic microorganisms. We functionalized a genetically engineered multimeric protein with both a targeting moiety (biotin) and either a fluorophore or a photosensitizer (SnCe6). Biofilm microcolonies of Aggregatibacter actinomycetemcomitans, a periodontal pathogen, were targeted with the multifunctional dodecamer. Streptavidin was used to couple biotinylated dodecamer to a biotinylated anti-A. actinomycetemcomitans antibody. This modular targeting approach enabled us to increase the loading of photosensitizer onto the cells by a cycle of amplification. Scanning laser confocal microscopy was used to characterize transport of fluorescently tagged dodecamer into the microcolonies and targeting of the cells with biotin-labeled, fluorescently tagged dodecamer. Light-induced activity of the targeted photosensitizer reduced the viability of A. actinomycetemcomitans biofilm, as indicated by membrane permeability to propidium iodide. The functionalized multimeric protein promises to be a useful tool for controlling periodontal biofilm consortia and offers a modular design whereby moieties that target different species can be readily combined with the functionalized protein construct.

Caspase-1 inhibitor reduced the lung injury in a mouse model of pleuropneumonia caused by Actinobacillus pleuropneumoniae.

The pathogenesis of porcine contagious pleuropneumonia is poorly understood. In the present study, a mouse model of intranasal infection by Actinobacillus pleuropneumoniae (App) was used to examine lung inflammation. The pathogical results of lung tissues showed that App-infected mice showed dyspnea and anorexia, with severe damage by acute hemorrhage, and infiltration of eosinophils and lymphocytes, as well as increased expression of caspase-1 p20, interleukin (IL)-1ß, IL-6, IL-8, IL-18 and tumor necrosis factor (TNF)-α. Caspase-1 inhibitors reduced both lung tissue damage and the expression of caspase-1 p20, IL-1ß, IL-6, IL-8, TNF-α and IL-18 in infected mice. These findings suggest that the caspase-1 dependent pyroptosis involved in the pathogenesis of the mouse pleuropneumonia caused by App and the inhibition of caspase-1 reduced the lung injury of this pleuropneumonia.

Efficacy of enteric-coated tilmicosin granules in pigs artificially infected with Actinobacillus pleuropneumoniae serotype 2.

BACKGROUND: Porcine infectious pleuropneumonia caused by Actinobacillus pleuropneumoniae (App) is one of the most serious infectious diseases in pigs and has brought huge economic losses to the world pig industry. The aim of this trial was to evaluate the effect of enteric-coated tilmicosin granule in the treatment and control of artificial infection of App. METHODS: Sixty Duroc and Yorkshire crossbred pigs (50 of which were artificially infected) were divided into six groups: BCG (Blank control group), ICG (Infection-only control group), HDG (High-dose enteric-coated tilmicosin granules), MDG (Medium-dose enteric-coated tilmicosin granules), LDG (Low-dose enteric-coated tilmicosin granules) and TPG (Tilmicosin premix drug control group). The cure rate, mortality, clinical respiratory score, body temperature score, weight gain, lung score and so on were recorded. RESULTS: The cure rate of HDG and MDG was as high as 90%, the mortality was 10%, and the clinical signs recovered quickly. CONCLUSION: The results showed that enteric-coated tilmicosin granules had obvious therapeutic effect on artificial infection, which could reduce the damage caused by the disease and reduce the mortality.