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Canadian Journal of Veterinary Research... Jul 2009A DNA microarray system was prepared and shown to facilitate identification and typing of Actinobacillus pleuropneumoniae. The DNA microarray, composed of 18 DNA...
A DNA microarray system was prepared and shown to facilitate identification and typing of Actinobacillus pleuropneumoniae. The DNA microarray, composed of 18 DNA polymerase chain reaction (PCR) amplicons printed on glass slides and arranged in 3 subarrays, was developed. These target DNA included 1 or multiple fragments of the outer membrane lipoprotein, apx toxin, capsular polysaccharide, and disulfide bound formation protein E (dsbE)-like genes of A. pleuropneumoniae. These arrayed target DNA retained their expected hybridization properties. The hybridization signal intensities ranged from the least-intense to the most-intense, 4626 to 9789 arbitrary fluorescence units, respectively. Cy3-probes of A. pleuropneumoniae strains labeled with multiplex PCR were hybridized to the DNA microarray. A total of 51 different A. pleuropneumoniae strains representing serotype 1 to 12 reference strains and clinical isolates were detected and typed by the DNA microarray. Twelve reference serotypes produced 11 distinct target DNA hybridization patterns, and hybridization patterns of serotypes 1 (n = 7), 3 (n = 5), and 7 (n = 6) field isolates were identical to hybridization patterns of reference serotypes 1, 3, and 7, respectively. Non-serotyped isolates 4, 6, and 11 (out of 21) from diseased pigs had identical hybridization patterns to reference serotypes 3, 7, and 1, respectively. The results show that the DNA microarray system described in the present study is a valuable tool for identifying and typing reference strains and isolates of A. pleuropneumoniae, and enables relatively rapid identification of non-serotyped isolates.
Topics: Actinobacillus Infections; Actinobacillus pleuropneumoniae; Animals; DNA, Bacterial; Oligonucleotide Array Sequence Analysis; Serotyping; Swine; Swine Diseases
PubMed: 19794891
DOI: No ID Found -
The Journal of Antimicrobial... Jan 2018To characterize ICEApl2, an SXT-related integrative and conjugative element (ICE) found in a clinical isolate of the porcine pathogen Actinobacillus pleuropneumoniae,...
Characterization of the Actinobacillus pleuropneumoniae SXT-related integrative and conjugative element ICEApl2 and analysis of the encoded FloR protein: hydrophobic residues in transmembrane domains contribute dynamically to florfenicol and chloramphenicol efflux.
OBJECTIVES
To characterize ICEApl2, an SXT-related integrative and conjugative element (ICE) found in a clinical isolate of the porcine pathogen Actinobacillus pleuropneumoniae, and analyse the functional nature of the encoded FloR.
METHODS
ICEApl2 was identified in the genome of A. pleuropneumoniae MIDG3553. Functional analysis was done using conjugal transfer experiments. MIDG3553 was tested for susceptibility to the antimicrobials for which resistance genes are present in ICEApl2. Lack of florfenicol/chloramphenicol resistance conferred by the encoded FloR protein was investigated by cloning and site-directed mutagenesis experiments in Escherichia coli.
RESULTS
ICEApl2 is 92660 bp and contains 89 genes. Comparative sequence analysis indicated that ICEApl2 is a member of the SXT/R391 ICE family. Conjugation experiments showed that, although ICEApl2 is capable of excision from the chromosome, it is not self-transmissible. ICEApl2 encodes the antimicrobial resistance genes floR, strAB, sul2 and dfrA1, and MIDG3553 is resistant to streptomycin, sulfisoxazole and trimethoprim, but not florfenicol or chloramphenicol. Cloning and site-directed mutagenesis of the floR gene revealed the importance of the nature of the hydrophobic amino acid residues at positions 160 and 228 in FloR for determining resistance to florfenicol and chloramphenicol.
CONCLUSIONS
Our results indicate that the nature of hydrophobic residues at positions 160 and 228 of FloR contribute dynamically to specific efflux of florfenicol and chloramphenicol, although some differences in resistance levels may depend on the bacterial host species. This is also, to our knowledge, the first description of an SXT/R391 ICE in A. pleuropneumoniae or any member of the Pasteurellaceae.
Topics: Actinobacillus pleuropneumoniae; Animals; Bacterial Proteins; Biological Transport; Chloramphenicol; Conjugation, Genetic; Drug Resistance, Multiple, Bacterial; Hydrophobic and Hydrophilic Interactions; Interspersed Repetitive Sequences; Microbial Sensitivity Tests; Pneumonia; Swine; Swine Diseases; Thiamphenicol
PubMed: 29029160
DOI: 10.1093/jac/dkx342 -
The Journal of Veterinary Medical... Sep 2019Actinobacillus species are known to be pathogenic to horses. To clarify etiological agents of actinobacillosis in Japanese adult horses, 27 isolates from Japanese...
Actinobacillus species are known to be pathogenic to horses. To clarify etiological agents of actinobacillosis in Japanese adult horses, 27 isolates from Japanese Thoroughbred racehorses putatively identified by matrix-assisted laser desorption ionization time-of-flight mass spectrometry as Actinobacillus were further identified by PCR of the A. equuli toxin gene, by CAMP test, and by 16S rRNA sequencing analysis. Actinobacillus equuli subsp. haemolyticus was isolated most frequently (16/27) and was related to respiratory infections. Actinobacillus equuli subsp. equuli (4/27) was isolated from chronic cases or concomitant with other bacterial infections. The remainder were A. pleuropneumoniae, unclassified Actinobacillus species and Pasteurella caballi. Actinobacillus equuli including subsp. haemolyticus and subsp. equuli were the species most frequently isolated from equine actinobacillosis in Japan.
Topics: Actinobacillus; Actinobacillus Infections; Animals; Bacterial Proteins; DNA, Bacterial; Horse Diseases; Horses; Japan; Pasteurella; Polymerase Chain Reaction; RNA, Ribosomal, 16S; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
PubMed: 31292334
DOI: 10.1292/jvms.19-0192 -
Journal of Bacteriology Mar 2024is an important respiratory pathogen that can cause porcine contagious pleuropneumonia (PCP), resulting in significant economic losses in swine industry. Microorganisms...
is an important respiratory pathogen that can cause porcine contagious pleuropneumonia (PCP), resulting in significant economic losses in swine industry. Microorganisms are subjected to drastic changes in environmental osmolarity. In order to alleviate the drastic rise or fall of osmolarity, cells activate mechanosensitive channels MscL and MscS through tension changes. MscL not only regulates osmotic pressure but also has been reported to secrete protein and uptake aminoglycoside antibiotic. However, MscL and MscS, as the most common mechanosensitive channels, have not been characterized in . In this study, the osmotic shock assay showed that MscL increased sodium adaptation by regulating cell length. The results of MIC showed that deletion of decreased the sensitivity of to multiple antibiotics, while deletion of rendered hypersensitive to penicillin. Biofilm assay demonstrated that MscL contributed the biofilm formation but MscS did not. The results of animal assay showed that MscL and MscS did not affect virulence . In conclusion, MscL is essential for sodium hyperosmotic tolerance, biofilm formation, and resistance to chloramphenicol, erythromycin, penicillin, and oxacillin. On the other hand, MscS is only involved in oxacillin resistance.IMPORTANCEBacterial resistance to the external environment is a critical function that ensures the normal growth of bacteria. MscL and MscS play crucial roles in responding to changes in both external and internal environments. However, the function of MscL and MscS in has not yet been reported. Our study shows that MscL plays a significant role in osmotic adaptation, antibiotic resistance, and biofilm formation of , while MscS only plays a role in antibiotic resistance. Our findings provide new insights into the functional characteristics of MscL and MscS in . MscL and MscS play a role in antibiotic resistance and contribute to the development of antibiotics for .
Topics: Animals; Swine; Actinobacillus pleuropneumoniae; Anti-Bacterial Agents; Virulence; Oxacillin; Sodium; Swine Diseases
PubMed: 38391161
DOI: 10.1128/jb.00429-23 -
BMC Veterinary Research Jul 2022Flagellin elicits potent immune response and may serve as a vaccine adjuvant. We previously reported that the N-terminus of flagellin (residues 1-99, nFliC) is...
BACKGROUND
Flagellin elicits potent immune response and may serve as a vaccine adjuvant. We previously reported that the N-terminus of flagellin (residues 1-99, nFliC) is sufficient for vaccine efficacy enhancement against Pasteurella multocida challenge in chickens. In this study, we futher tested the adjuvancy of nFliC in a subunit vaccine against the pig pathogen Actinobacillus pleuropneumoniae in a mice model. For vaccine formulation, the antigen ApxIIPF (the pore-forming region of the exotoxin ApxII) was combined with nFliC, either through genetic fusion or simple admixture.
RESULTS
Immune analysis showed that nFliC, introduced through genetic fusion or admixture, enhanced both humoral (antibody levels) and cellular (T cell response and cytokine production) immunity. In a challenge test, nFliC increased vaccine protective efficacy to 60-80%, vs. 20% for the antigen-only group. Further analysis showed that, even without a supplemental adjuvant such as mineral salt or oil emulsion, genetically linked nFliC still provided significant immune enhancement.
CONCLUSIONS
We conclude that nFliC is a versatile and potent adjuvant for vaccine formulation.
Topics: Actinobacillus Infections; Actinobacillus pleuropneumoniae; Animals; Antibodies, Bacterial; Bacterial Vaccines; Chickens; Flagellin; Mice; Swine; Swine Diseases; Vaccine Efficacy
PubMed: 35842618
DOI: 10.1186/s12917-022-03380-8 -
Acta Veterinaria Scandinavica 2002The objective of the study was to identify risk factors for reintroduction of Actinobacillus pleuopneumoniae and Mycoplasma hyopneumoniae (enzootic pneumonia) onto pig...
Incidence of reinfections with Mycoplasma hyopneumoniae and Actinobacillus pleuropneumoniae in pig farms located in respiratory-disease-free regions of Switzerland--identification and quantification of risk factors.
The objective of the study was to identify risk factors for reintroduction of Actinobacillus pleuopneumoniae and Mycoplasma hyopneumoniae (enzootic pneumonia) onto pig farms in areas in Switzerland that were involved in an eradication programme from 1996 to 1999 and to assess the role of dealers in relation to these reinfections. The study was based on the comparison of pig farms that were reinfected in the year 2000 (cases) and pig farms that remained uninfected in the same area (controls). Additionally, data were collected from Swiss pig dealers and transport companies. Out of a total of 3983 farms, 107 farms were reinfected in the year 2000. The incidences were 0.1% for Actinobacillus pleuopneumoniae and 2.6% for Mycoplasma hyopneumoniae (enzootic pneumonia). Compared to reinfection rates prior to the eradication programme, this is a considerable reduction. Statistically significant risk factors for the reinfection were 'finishing farm', 'large mixed breeding-finishing farm', 'reinfected neighbour' and 'parking site for pig transport vehicles close to the farm'. Pig farmers that purchased pigs from only one supplier per batch had a lower risk of reintroducing infection (protective factor). As long as infected and uninfected regions co-exist in Switzerland, direct and indirect contact between farms, pig herds and slaughter sites via transport vehicles are a major pathway of disease spread. Risk management measures linked to these contacts are therefore of key importance. The survey of dealers indicated various areas for improvement such as strategic planning of pick-up routes or cleaning and disinfecting of trucks.
Topics: Actinobacillus Infections; Actinobacillus pleuropneumoniae; Animals; Incidence; Mycoplasma Infections; Recurrence; Risk Factors; Surveys and Questionnaires; Swine; Swine Diseases; Switzerland
PubMed: 12564544
DOI: 10.1186/1751-0147-43-145 -
American Journal of Physiology.... Apr 2009Urea transporters (UTs) effect rapid flux of urea across biological membranes. In the mammalian kidney, UT activity is essential for effective urine concentration. In...
Urea transporters (UTs) effect rapid flux of urea across biological membranes. In the mammalian kidney, UT activity is essential for effective urine concentration. In bacteria, UT-mediated urea uptake permits intracellular urease to degrade urea to ammonia and CO(2), a process that either buffers acid loads or provides nutrient nitrogen. We have characterized the urea transport channel protein ApUT from Actinobacillus pleuropneumoniae. Kinetic analysis of bacterial inside-out membranes enriched in ApUT showed approximately 28-fold increase in urea permeability (3.3 +/- 0.4 x 10(-4) cm/s) compared with control vesicles (0.11 +/- 0.02 x 10(-4) cm/s). In addition to urea, ApUT also conducts water. Urea and water transport across the channel was phloretin and mercury inhibitable, and the site of inhibition may be located on the cytoplasmic side of the protein. Glycerol and urea analogs, such as methylamine, dimethylurea, formamide, acetamide, methylurea, propanamide, and ethylamine did not permeate across ApUT.
Topics: Actinobacillus pleuropneumoniae; Bacterial Proteins; Biological Transport; Cell Membrane Permeability; Cloning, Molecular; Escherichia coli; Glycerol; Kinetics; Membrane Transport Proteins; Mercuric Chloride; Phloretin; Urea; Water
PubMed: 19144751
DOI: 10.1152/ajpregu.90726.2008 -
Journal of Bacteriology Jan 2003Actinobacillus pleuropneumoniae has been considered nonmotile and nonflagellate. In this work, it is demonstrated that A. pleuropneumoniae produces flagella composed of...
Actinobacillus pleuropneumoniae has been considered nonmotile and nonflagellate. In this work, it is demonstrated that A. pleuropneumoniae produces flagella composed of a 65-kDa protein with an N-terminal amino acid sequence that shows 100% identity with those of Escherichia coli, Salmonella, and Shigella flagellins. The DNA sequence obtained through PCR of the fliC gene in A. pleuropneumoniae showed considerable identity (93%) in its 5' and 3' ends with the DNA sequences of corresponding genes in E. coli, Salmonella enterica, and Shigella spp. The motility of A. pleuropneumoniae was observed in tryptic soy or brain heart infusion soft agar media, and it is influenced by temperature. Flagella and motility may be involved in the survival and pathogenesis of A. pleuropneumoniae in pigs.
Topics: Actinobacillus Infections; Actinobacillus pleuropneumoniae; Animals; Culture Media; Flagella; Flagellin; Microscopy, Electron; Molecular Sequence Data; Movement; Sequence Analysis, DNA; Swine; Swine Diseases
PubMed: 12511514
DOI: 10.1128/JB.185.2.664-668.2003 -
International Journal of Molecular... Jul 2023(APP) is the causative pathogen of porcine pleuropneumonia, a highly contagious respiratory disease in the pig industry. The increasingly severe antimicrobial...
(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 β-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.
Topics: Animals; Swine; Mice; Pleuropneumonia; Actinobacillus pleuropneumoniae; Toll-Like Receptor 4; Actinobacillus; Tight Junctions; Lung; Actinobacillus Infections; Mycoplasma Infections; Tea; Swine Diseases
PubMed: 37511601
DOI: 10.3390/ijms241411842 -
Journal of Comparative Pathology Jul 2022In-vivo models of Actinobacillus pleuropneumoniae (App) infection in pigs are required for the development of vaccines and investigations of pathogenicity. Existing...
In-vivo models of Actinobacillus pleuropneumoniae (App) infection in pigs are required for the development of vaccines and investigations of pathogenicity. Existing models cause severe respiratory disease with pulmonary oedema, dyspnoea and severe thoracic pain, and careful monitoring and early intervention with euthanasia is, therefore, needed to avoid unnecessary suffering in experimental animals. As a potential replacement for the existing respiratory infection model, an in-vivo protocol was evaluated using intradermal or subcutaneous injection of different App strains and Apx toxins into the abdominal skin of pigs. High concentrations of serovar 1 and serovar 10 App induced diffuse visible dermal oedema and inflammation. Injection of Apx toxins alone did not adequately produce macroscopic lesions, although an influx of inflammatory cells was seen on histopathology. ApxI-producing strains of App induced more inflammation than ApxII- and ApxIII-producing strains. Induction of skin lesions by injection of App or Apx toxins was not sufficiently repeatable or discrete for a robust experimental model that could be used for assessment of novel interventions.
Topics: Actinobacillus Infections; Actinobacillus pleuropneumoniae; Animals; Bacterial Proteins; Bacterial Toxins; Edema; Hemolysin Proteins; Inflammation; Models, Theoretical; Swine; Swine Diseases
PubMed: 35817536
DOI: 10.1016/j.jcpa.2022.04.004