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.
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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. (+info)
Expression of cyclooxygenase-2 in swine naturally infected with Actinobacillus pleuropneumoniae.
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Cyclooxygenase-2 (COX-2) was detected and localized in 15 pigs with naturally occurring pleuropneumonia using a 437-base pair digoxigenin-labeled cDNA probe in an in situ hybridization protocol. Histopathologic changes in the acute stage were characterized by coagulative necrosis of lung parenchyma, hemorrhage, vascular thrombosis, edema, fibrin deposition, and infiltration of lung parenchyma by neutrophils and alveolar macrophages in nine pigs. In chronic lesions, a thick layer of granulation tissue surrounded foci of pulmonary necrosis in six pigs. All 15 pigs infected with Actinobacillus pleuropneumoniae, confirmed by bacterial isolation, had distinct positive hybridization signals for COX-2 in bronchial, bronchiolar epithelial cells, alveolar macrophages, neutrophils, and type I pneumocytes. COX-2 expression was detected primarily in neutrophils from pigs with acute lesions and primarily in alveolar macrophages from pigs with chronic lesions. The results suggest that a prostanoid product of COX-2 is an important component of the inflammatory response to acute and chronic A. pleuropneumoniae infection. (+info)
Prevalence of cytolethal distending toxin production in periodontopathogenic bacteria.
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Cytolethal distending toxin (CDT) is a newly identified virulence factor produced by several pathogenic bacteria implicated in chronic infection. Seventy three strains of periodontopathogenic bacteria were examined for the production of CDT by a HeLa cell bioassay and for the presence of the cdt gene by PCR with degenerative oligonucleotide primers, which were designed based on various regions of the Escherichia coli and Campylobacter cdtB genes, which have been successfully used for the identification and cloning of cdtABC genes from Actinobacillus actinomycetemcomitans Y4 (M. Sugai et al., Infect. Immun. 66:5008-5019, 1998). CDT activity was found in culture supernatants of 40 of 45 tested A. actinomycetemcomintans strains, but the titer of the toxin varied considerably among these strains. PCR experiments indicated the presence of Y4-type cdt sequences in these strains, but the rest of A. actinomycetemcomitans were negative by PCR amplification and also by Southern blot analysis for the cdtABC gene. In the 40 CDT-positive strains, Southern hybridization with HindIII-digested genomic DNA revealed that there are at least 6 restriction fragment length polymorphism types. This suggests that the cdtABC flanking region is highly polymorphic, which may partly explain the variability of the CDT activity in the culture supernatants. The rest of tested strains of periodontopathogenic bacteria did not have detectable CDT production by the HeLa cell assay and for cdtB sequences by PCR analysis under our experimental conditions. These results strongly suggested that CDT is a unique toxin predominantly produced by A. actinomycetemcomitans among periodontopathogenic bacteria. (+info)
Evidence of nitric oxide synthase 2 activity in swine naturally infected with Actinobacillus pleuropneumoniae.
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Evidence of nitric oxide synthase (NOS) 2 activity was determined by formation of nitrotyrosine (a reaction product of peroxynitrite) and by activation of poly(ADP-ribose) synthetase (PARS) in NOS2-expressed pleuropneumonic lungs from 20 pigs naturally infected with Actinobacillus pleuropneumoniae using immunohistochemistry. Intense immunostaining for nitrotyrosine residue was seen within the lung lesions from A. pleuropneumoniae-infected pigs, but it was minimal in the unaffected parts of the lung from A. pleuropneumoniae-infected pigs and in the normal lung from control pigs. Staining was especially strong in neutrophils and macrophages in the periphery of the lesions and within the alveolar spaces. There was close cell-to-cell correlation when serial sections were examined by immunohistochemistry for NOS2 and nitrotyrosine in each of the 20 lung samples. Expression of PARS was always present within inflammatory lesions but was minimal in the unaffected lung of A. pleuropneumoniae-infected pigs. Macrophages in alveolar spaces frequently exhibited strong staining for PARS. Colocalization of nitrotyrosine and PARS antigen was especially prominent in macrophages in the periphery of lesions. NOS2 expression in pleuropneumonic areas associated with protein nitrosation and PARS suggests that NOS2 is functionally active during infections caused by A. pleuropneumoniae. (+info)
Descriptive epidemiology of late-term abortions associated with the mare reproductive loss syndrome in central Kentucky.
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Epidemiological and pathological findings of 433 late-term abortions associated with the mare reproductive loss syndrome (MRLS) in central Kentucky were identified by reviewing the records of the University of Kentucky Livestock Diseases Diagnostic Center. The distribution of dates of abortion was clustered during a brief period of time, presumably from a simultaneous environmental exposure. The most common pathological findings were microscopic pulmonary lesions consisting of squamous epithelial cells present in alveoli with or without concurrent infiltration of inflammatory cells (neutrophils, macrophages, or monocytes) in the interstitium or within alveoli. Isolation of a non-beta-hemolytic Streptococcus (52% of fetuses) or an Actinobacillus sp. (19% of fetuses) was common. Placentitis or funisitis was identified in 44% of fetuses. No single pathological finding, however, was pathognomonic for MRLS-associated late-term abortion. This report describes the pathological findings characterizing the MRLS-associated abortion. A cause of MRLS could not be determined from necropsy findings. (+info)
Tight-adherence genes of Actinobacillus actinomycetemcomitans are required for virulence in a rat model.
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Actinobacillus actinomycetemcomitans is a Gram-negative coccobacillus that has been associated with localized aggressive periodontitis and infections of the heart, brain, and urinary tract. Wild-type clinical isolates have the remarkable ability to adhere tenaciously and nonspecifically to solid surfaces such as glass, plastic, and hydroxyapatite. Adherence by A. actinomycetemcomitans is mediated by the tight-adherence (tad) gene locus, which consists of 14 genes (flp-1-flp-2-tadV-rcpCAB-tadZABCDEFG). All but 2 of the genes have been shown to be required for the secretion and assembly of long, bundled Flp1 fibrils. To test whether the tad locus is required for colonization and disease, we developed a rat model for periodontal disease. To mimic the natural route of infection, Sprague-Dawley rats were inoculated orally by adding bacteria directly to their food for 8 days. After inoculation with wild-type or mutant strains defective in adherence (flp-1 and tadA), the rats were assessed for colonization of the oral cavity and pathogenesis. Wild-type A. actinomycetemcomitans was able to colonize and persist for at least 12 weeks in the oral cavity, elicit a humoral immune response, and cause significant bone loss in rats. In contrast, rats fed flp-1 or tadA mutant strains showed no bone loss and their immune responses were indistinguishable from those of the uninoculated controls. These results demonstrate the critical importance of the tad locus in the colonization and pathogenesis of A. actinomycetemcomitans. (+info)
Identification of Actinobacillus pleuropneumoniae genes important for survival during infection in its natural host.
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Actinobacillus pleuropneumoniae is a strict respiratory tract pathogen of swine and is the causative agent of porcine pleuropneumonia. We have used signature-tagged mutagenesis (STM) to identify genes required for survival of the organism within the pig. A total of 2,064 signature-tagged Tn10 transposon mutants were assembled into pools of 48 each, and used to inoculate pigs by the endotracheal route. Out of 105 mutants that were consistently attenuated in vivo, only 11 mutants showed a >2-fold reduction in growth in vitro compared to the wild type, whereas 8 of 14 mutants tested showed significant levels of attenuation in pig as evidenced from competitive index experiments. Inverse PCR was used to generate DNA sequence of the chromosomal domains flanking each transposon insertion. Only one sibling pair of mutants was identified, but three apparent transposon insertion hot spots were found--an anticipated consequence of the use of a Tn10-based system. Transposon insertions were found within 55 different loci, and similarity (BLAST) searching identified possible analogues or homologues for all but four of these. Matches included proteins putatively involved in metabolism and transport of various nutrients or unknown substances, in stress responses, in gene regulation, and in the production of cell surface components. Ten of the sequences have homology with genes involved in lipopolysaccharide and capsule production. The results highlight the importance of genes involved in energy metabolism, nutrient uptake and stress responses for the survival of A. pleuropneumoniae in its natural host: the pig. (+info)
Caspase 1 involvement in human monocyte lysis induced by Actinobacillus actinomycetemcomitans leukotoxin.
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Actinobacillus actinomycetemcomitans, an oral bacterium implicated in the etiology of periodontal diseases, produces a leukotoxin that selectively lyses primate neutrophils and monocytes, the major populations of defense cells in the periodontium. Though lysis requires expression of the receptor lymphocyte function-associated molecule 1 (LFA-1) on the cell surface, not all LFA-1-expressing leukocyte populations are equally susceptible to the toxin. In this study, the susceptibility of human leukocytes to leukotoxin-induced lysis is compared to their expression of LFA-1 and the activity of caspase 1. Cytolysis was determined by the activity of lactate dehydrogenase released from peripheral human leukocytes after 1-h exposure to leukotoxin. Monocytes were lysed at leukotoxin concentrations of > or = 5 ng/ml, while the corresponding values for neutrophils and lymphocytes were approximately 10 times greater. Similar LFA-1 expression was found in all susceptible cell populations irrespective of their degree of sensitivity to the toxin. Exposure of monocytes to leukotoxin increased their caspase 1 activity about fivefold within 10 to 20 min. Presence of the caspase 1 inhibitor Ac-YVAD-CMK significantly blocked the leukotoxin-induced lysis of monocytes only. At sublytic concentrations, leukotoxin induced no apoptotic activity in monocytes, as revealed by the lack of caspase 3 activation and DNA fragmentation. Monocytes are the most lysis-sensitive leukocytes for A. actinomycetemcomitans leukotoxin. Their lysis by this toxin depends on caspase 1 activation and proceeds through a process that differs from classical apoptosis. (+info)