Identification of Novel Zoonotic Activity of Bartonella spp., France.

Certain Bartonella species are known to cause afebrile bacteremia in humans and other mammals, including B. quintana, the agent of trench fever, and B. henselae, the agent of cat scratch disease. Reports have indicated that animal-associated Bartonella species may cause paucisymptomatic bacteremia and endocarditis in humans. We identified potentially zoonotic strains from 6 Bartonella species in samples from patients who had chronic, subjective symptoms and who reported tick bites. Three strains were B. henselae and 3 were from other animal-associated Bartonella spp. (B. doshiae, B. schoenbuchensis, and B. tribocorum). Genomic analysis of the isolated strains revealed differences from previously sequenced Bartonella strains. Our investigation identifed 3 novel Bartonella spp. strains with human pathogenic potential and showed that Bartonella spp. may be the cause of undifferentiated chronic illness in humans who have been bitten by ticks.

Aspergillus fumigatus in Poultry.

Aspergillus fumigatus remains a major respiratory pathogen in birds. In poultry, infection by A. fumigatus may induce significant economic losses particularly in turkey production. A. fumigatus develops and sporulates easily in poor quality bedding or contaminated feedstuffs in indoor farm environments. Inadequate ventilation and dusty conditions increase the risk of bird exposure to aerosolized spores. Acute cases are seen in young animals following inhalation of spores, causing high morbidity and mortality. The chronic form affects older birds and looks more sporadic. The respiratory tract is the primary site of A. fumigatus development leading to severe respiratory distress and associated granulomatous airsacculitis and pneumonia. Treatments for infected poultry are nonexistent; therefore, prevention is the only way to protect poultry. Development of avian models of aspergillosis may improve our understanding of its pathogenesis, which remains poorly understood.

Inducible expression of beta defensins by human respiratory epithelial cells exposed to Aspergillus fumigatus organisms.

BACKGROUND: Aspergillus fumigatus, a saprophytic mould, is responsible for life-threatening, invasive pulmonary diseases in immunocompromised hosts. The role of the airway epithelium involves a complex interaction with the inhaled pathogen. Antimicrobial peptides with direct antifungal and chemotactic activities may boost antifungal immune response. RESULTS: The inducible expression of defensins by human bronchial epithelial 16HBE cells and A549 pneumocyte cells exposed to A. fumigatus was investigated. Using RT-PCR and real time PCR, we showed an activation of hBD2 and hBD9 defensin genes: the expression was higher in cells exposed to swollen conidia (SC), compared to resting conidia (RC) or hyphal fragments (HF). The kinetics of defensin expression was different for each one, evoking a putative distinct function for each investigated defensin. The decrease of defensin expression in the presence of heat-inactivated serum indicated a possible link between defensins and the proteins of the host complement system. The presence of defensin peptide hBD2 was revealed using immunofluorescence that showed a punctual cytoplasmic and perinuclear staining. Quantification of the cells stained with anti hBD2 antibody demonstrated that SC induced a greater number of cells that synthesized hBD2, compared to RC or HF. Labelling of the cells with anti-hBD-2 antibody showed a positive immunofluorescence signal around RC or SC in contrast to HF. This suggests co-localisation of hBD2 and digested conidia. The HBD2 level was highest in the supernatants of cells exposed to SC, as was determined by sandwich ELISA. Experiments using neutralising anti-interleukine-1beta antibody reflect the autocrine mechanism of defensin expression induced by SC. Investigation of defensin expression at transcriptional and post-transcriptional levels demonstrated the requirement of transcription as well as new protein synthesis during A. fumigatus defensin induction. Finally, induced defensin expression in primary culture of human respiratory cells exposed to A. fumigatus points to the biological significance of described phenomena. CONCLUSION: Our findings provide evidence that respiratory epithelium might play an important role in the immune response during Aspergillus infection. Understanding the mechanisms of regulation of defensin expression may thus lead to new approaches that could enhance expression of antimicrobial peptides for potential therapeutic use during aspergillosis treatment.

Clinical, mycological and pathological findings in turkeys experimentally infected by Aspergillus fumigatus.

Experimental aspergillosis was induced in 1-day-old turkeys by intra-air-sac inoculation of a spore suspension of a 3-day-old Aspergillus fumigatus culture (CBS 144.89) containing 10(7) spores. Ten additional poults were used as controls. Infected and non-infected animals were closely observed at least twice a day for the appearance of clinical signs and were sequentially sacrificed at days 1, 2, 3, 5 and 7 post-inoculation. In the infected group, most lung tissues and air sac swabs were culture positive from day 1 to day 5. At 1 day post-inoculation, air sac membranes were multifocally and moderately to severely thickened by an oedema and covered by an exudate. A small number of germinating conidia were present in the superficial exudate, already giving rise to small radiating hyphae. Lung lesions were mild, dominated by a diffuse congestion and a mild heterophilic infiltration. From 2 to 3 days post-inoculation, air sac membranes were more severely affected and several granulomas were observed. Both granulomas and exudates were rich in germinated conidia and hyphae. Pulmonary lesions consisted in a diffuse pneumonia. Five days post-inoculation, air sac membrane lesions progressed to a severe, multifocal, heterophilic and granulomatous inflammation. Seven days post-inoculation, a reduction of the severity of the diffuse pneumonia was detected. Concomitantly, the fungal elements were mainly observed as fragmented tubules in the cytoplasm of multinucleate giant cells. The present study demonstrated that healthy turkey poults might be able to withstand exposure to 10(7) A. fumigatus spores.

Aspergillus fumigatus conidia inhibit tumour necrosis factor- or staurosporine-induced apoptosis in epithelial cells.

A major innate immune response to inhaled conidia of the opportunistic pathogen Aspergillus fumigatus (Af) is the synthesis of pro-inflammatory cytokines, which include tumour necrosis factor (TNF)-alpha, a known inducer of apoptosis. Modulation of host cell apoptosis has been reported to be one of the mechanisms whereby pathogens overcome host cell defences. Our study was designed to investigate whether or not Af conidia could modulate apoptosis induced by TNF-alpha or staurosporine (STS). Exposure of epithelial cells treated by these inducers and exposed to Af conidia decreased the number of apoptotic cells detected by Annexin V staining, analysis of nuclear morphology, terminal deoxynucleotidyl transferase-mediated fluorescein-dUTP nick end-labelling reaction and immunoblotting. Inhibition of apoptosis by Af conidia was seen in cells of the A549 pneumocyte II line, human tracheal epithelial 16HBE and primary human respiratory cells. Inhibition of apoptosis by Af conidia was also observed when apoptosis was induced by co-cultivating A549 cells with activated human alveolar macrophages. Unlike Af conidia, conidia of Cladosporium cladosporioides as well as latex beads or killed Af conidia have no inhibitory effect on TNF-alpha or STS-induced apoptosis. For TNF-induced apoptosis, the observed anti-apoptotic effect of Af conidia was found to be associated with a significant reduction of caspase-3.

Establishment and characterization of continuous hematopoietic progenitors-derived pig normal mast cell lines.

Mast cells (MCs) are tissue resident, hematopoietic stem cells-derived elements, distributed throughout the body. They are the pivotal mediating cells of allergic reactions. In addition, in mice, MCs play a critical role in the defense against several pathogens, such as bacteria, parasites and viruses. Whereas the biology of rodent and human MCs has been extensively studied using in vitro derived populations, the role of MCs in pigs has not yet been evaluated, given the very low availability of pure porcine MCs populations. In the present report, we describe an original method to obtain continuous factor-dependent normal pig MCs (PMC) lines from fetal hematopoietic progenitors. These Stem Cell Factor (SCF) and Interleukin-3- (IL-3)-dependent PMC lines retain their capacity to growth after conventional freezing methods and exhibit most of the morphological and biochemical properties of normal, although immature, MCs, including metachromatic granules containing sulfated polysaccharides, the expression of c-kit and high-affinity IgE receptors (FcepsilonRI), and the ability to store histamine that is released upon cross-linking of FcepsilonRI. In vitro derived PMC lines might thus be valuable tools to further investigate the reactivity of these elements towards several parasites frequently encountered in pig, such as, but not limited to, Ascaris suum, Trichinella spiralis or Trichuris suis, or towards antigens derived from these pathogens.

Lymphoid activation: a confounding factor in AIDS vaccine development?

In a previous vaccination trial, inoculation of env gene DNA failed to elicit a detectable antibody response, yet accelerated virus dissemination in most immunized cats following challenge with feline immunodeficiency virus. This result raised the possibility that cell-mediated immune responses had given rise to immune-mediated enhancement of infection. Since high-level replication of immunodeficiency viruses in lymphocytes requires cellular activation, antigen-specific responses or non-specific polyclonal activation may have increased the frequency of optimal target cells. In the present DNA vaccination trial, although designed so as to minimize non-specific polyclonal activation, immune-mediated enhancement was nonetheless observed in certain immunized cats. Moreover, rapid virus dissemination in vivo was associated with the presence of T-helper responses prior to challenge, and was linked to increased susceptibility of lymphocytes to ex vivo infection. Immune activation may thus be a confounding factor in vaccination against lentivirus infection, diminishing vaccine efficacy and giving rise to immune-mediated enhancement.