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Melanie Brutyn,
Katharina D'Herde,
Maarten Dhaenens,
Pascale Van Rooij,
Elin Verbrugghe,
Alex D Hyatt,
Siska Croubels, Dieter Deforce,
Richard Ducatelle,
Freddy Haesebrouck,
An Martel,
Frank Pasmans
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ABSTRACT: Global amphibian declines are in part driven by the chytrid fungus Batrachochytrium dendrobatidis, causing superficial dermatomycosis with epidermal hyperplasia and hyperkeratosis in infected amphibians. The susceptibility to chytridiomycosis and the severity of epidermal lesions in amphibians with chytridiomycosis are not consistent across species or even among individuals. Severe infections cause death of the animal most likely through disturbance of ion homeostasis. The mechanism by which this superficial skin infection results in epidermal lesions has so far eluded precise definition. It was the aim of this study to unravel how B. dendrobatidis causes alterations that affect skin integrity. Exposure of Xenopus laevis skin to B. dendrobatidis zoospore supernatant using skin explants and Ussing chambers caused rapid disruption of intercellular junctions, demonstrated using histology and transmission electron microscopy. The loss of intercellular junctions led to detachment-induced cell apoptosis, or anoikis. The zoospore supernatant induced neither apoptosis nor necrosis in isolated primary keratinocytes of X. laevis. This supports the idea that the loss of cell contacts triggered apoptosis in the skin explants. Mass spectrometric analysis of the protein composition of the supernatant revealed a complex mixture, including several new virulence associated proteins, such as proteases, biofilm-associated proteins and a carotenoid ester lipase. Protease and lipase activity of the supernatant was confirmed with a protease and lipase assay. In conclusion, B. dendrobatidis zoospores produce a complex mixture of proteins that quickly disturbs epidermal intercellular junctions leading to anoikis in the anuran skin. The role of the identified proteins in this process remains to be determined.
Fungal Genetics and Biology 08/2012; 49(10):830-7. · 3.74 Impact Factor
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Elin Verbrugghe,
Virginie Vandenbroucke,
Maarten Dhaenens,
Neil Shearer,
Joline Goossens,
Sarah De Saeger,
Mia Eeckhout,
Katharina D'Herde,
Arthur Thompson, Dieter Deforce,
Filip Boyen,
Bregje Leyman,
Alexander Van Parys,
Patrick De Backer,
Freddy Haesebrouck,
Siska Croubels,
Frank Pasmans
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ABSTRACT: The mycotoxin T-2 toxin and Salmonella Typhimurium infections pose a significant threat to human and animal health. Interactions between both agents may result in a different outcome of the infection. Therefore, the aim of the presented study was to investigate the effects of low and relevant concentrations of T-2 toxin on the course of a Salmonella Typhimurium infection in pigs. We showed that the presence of 15 and 83 μg T-2 toxin per kg feed significantly decreased the amount of Salmonella Typhimurium bacteria present in the cecum contents, and a tendency to a reduced colonization of the jejunum, ileum, cecum, colon and colon contents was noticed. In vitro, proteomic analysis of porcine enterocytes revealed that a very low concentration of T-2 toxin (5 ng/mL) affects the protein expression of mitochondrial, endoplasmatic reticulum and cytoskeleton associated proteins, proteins involved in protein synthesis and folding, RNA synthesis, mitogen-activated protein kinase signaling and regulatory processes. Similarly low concentrations (1-100 ng/mL) promoted the susceptibility of porcine macrophages and intestinal epithelial cells to Salmonella Typhimurium invasion, in a SPI-1 independent manner. Furthermore, T-2 toxin (1-5 ng/mL) promoted the translocation of Salmonella Typhimurium over an intestinal porcine epithelial cell monolayer. Although these findings may seem in favour of Salmonella Typhimurium, microarray analysis showed that T-2 toxin (5 ng/mL) causes an intoxication of Salmonella Typhimurium, represented by a reduced motility and a downregulation of metabolic and Salmonella Pathogenicity Island 1 genes. This study demonstrates marked interactions of T-2 toxin with Salmonella Typhimurium pathogenesis, resulting in bacterial intoxication.
Veterinary Research 03/2012; 43(1):22. · 4.06 Impact Factor
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ABSTRACT: Ventilator-associated pneumonia (VAP) is a common nosocomial infection in mechanically ventilated patients. Biofilm formation is one of the mechanisms through which the endotracheal tube (ET) facilitates bacterial contamination of the lower airways. In the present study, we analyzed the composition of the ET biofilm flora by means of culture dependent and culture independent (16 S rRNA gene clone libraries and pyrosequencing) approaches. Overall, the microbial diversity was high and members of different phylogenetic lineages were detected (Actinobacteria, beta-Proteobacteria, Candida spp., Clostridia, epsilon-Proteobacteria, Firmicutes, Fusobacteria and gamma-Proteobacteria). Culture dependent analysis, based on the use of selective growth media and conventional microbiological tests, resulted in the identification of typical aerobic nosocomial pathogens which are known to play a role in the development of VAP, e.g. Staphylococcus aureus and Pseudomonas aeruginosa. Other opportunistic pathogens were also identified, including Staphylococcus epidermidis and Kocuria varians. In general, there was little correlation between the results obtained by sequencing 16 S rRNA gene clone libraries and by cultivation. Pyrosequencing of PCR amplified 16 S rRNA genes of four selected samples resulted in the identification of a much wider variety of bacteria. The results from the pyrosequencing analysis suggest that these four samples were dominated by members of the normal oral flora such as Prevotella spp., Peptostreptococcus spp. and lactic acid bacteria. A combination of methods is recommended to obtain a complete picture of the microbial diversity of the ET biofilm.
PLoS ONE 01/2012; 7(6):e38401. · 4.09 Impact Factor
