An Insertion Sequence-Dependent Plasmid Rearrangement in Aeromonas salmonicida Causes the Loss of the Type Three Secretion System

Aeromonas salmonicida, a bacterial fish pathogen, possesses a functional Type Three Secretion System (TTSS), which is essential for its virulence. The genes for this system are mainly located in a single region of the large pAsa5 plasmid. Bacteria lose the TTSS region from this plasmid through rearrangements when grown in stressful growth conditions. The A. salmonicida genome is rich in insertion sequences (ISs), which are mobile DNA elements that can cause DNA rearrangements in other bacterial species. pAsa5 possesses numerous ISs. Three IS11s from the IS256 family encircle the rearranged regions. To confirm that these IS11s are involved in pAsa5 rearrangements, 26 strains derived from strain A449 and two Canadian isolates (01-B526 and 01-B516) with a pAsa5 rearrangement were tested using a PCR approach to determine whether the rearrangements were the result of an IS11-dependent process. Nine out of the 26 strains had a positive PCR result, suggesting that the rearrangement in these strains were IS-dependent. The PCR analysis showed that all the rearrangements in the A449-derived strains were IS11-dependent process while the rearrangements in 01-B526 and 01-B516 could only be partially coupled to the action of IS11. Unidentified elements that affect IS-dependent rearrangements may be present in 01-B526 and 01-B516. Our results suggested that pAsa5 rearrangements involve IS11. This is the first study showing that ISs are involved in plasmid instability in A. salmonicida

Development of an In Vitro Model for the Multi-Parametric Quantification of the Cellular Interactions between Candida Yeasts and Phagocytes

We developed a new in vitro model for a multi-parameter characterization of the time course interaction of Candida fungal cells with J774 murine macrophages and human neutrophils, based on the use of combined microscopy, fluorometry, flow cytometry and viability assays. Using fluorochromes specific to phagocytes and yeasts, we could accurately quantify various parameters simultaneously in a single infection experiment: at the individual cell level, we measured the association of phagocytes to fungal cells and phagocyte survival, and monitored in parallel the overall phagocytosis process by measuring the part of ingested fungal cells among the total fungal biomass that changed over time. Candida albicans, C. glabrata, and C. lusitaniae were used as a proof of concept: they exhibited species-specific differences in their association rate with phagocytes. The fungal biomass uptaken by the phagocytes differed significantly according to the Candida species. The measure of the survival of fungal and immune cells during the interaction showed that C. albicans was the more aggressive yeast in vitro, destroying the vast majority of the phagocytes within five hours. All three species of Candida were able to survive and to escape macrophage phagocytosis either by the intraphagocytic yeast-to-hyphae transition (C. albicans) and the fungal cell multiplication until phagocytes burst (C. glabrata, C. lusitaniae), or by the avoidance of phagocytosis (C. lusitaniae). We demonstrated that our model was sensitive enough to quantify small variations of the parameters of the interaction. The method has been conceived to be amenable to the high-throughput screening of mutants in order to unravel the molecular mechanisms involved in the interaction between yeasts and host phagocytes

Use of in vivo induced antigen technology to identify genes from Aeromonas salmonicida subsp. salmonicida that are specifically expressed during infection of the rainbow trout Oncorhynchus mykiss

BACKGROUND: Aeromonas salmonicida is a major fish pathogen associated with mass mortalities in salmonid fish. In the present study, we applied In Vivo Induced Antigen Technology (IVIAT), a technique that relies on antibodies adsorbed against in vitro cultures of the pathogen, to a clinical isolate of A. salmonicida subsp. salmonicida. RESULTS: The results from IVIAT allowed identification of four proteins that were upregulated in the fish samples: A UDP-3-O-acyl-N-acetylglucosamine deacetylase, an RNA polymerase sigma factor D as well as TonB and a hypothetical protein. Subsequent investigations were performed using real-time PCR and cDNA synthesised from infected spleen, liver and anterior kidneys. These confirmed that the transcription level of each of these genes was significantly upregulated during the infection process compared to bacteria in vitro. CONCLUSIONS: The present studied identified four genes that were upregulated during the infectious process and are likely to play a role in the virulence of A. salmonicida. Because these are antigenic they might constitute potential targets for the development of new vaccine as well as therapeutic agents

Complement activation in the injured central nervous system: Another dual-edged sword?

The complement system, a major component of the innate immune system, is becoming increasingly recognised as a key participant in physiology and disease. The awareness that immunological mediators support various aspects of both normal central nervous system (CNS) function and pathology has led to a renaissance of complement research in neuroscience. Various studies have revealed particularly novel findings on the wide-ranging involvement of complement in neural development, synapse elimination and maturation of neural networks, as well as the progression of pathology in a range of chronic neurodegenerative disorders, and more recently, neurotraumatic events, where rapid disruption of neuronal homeostasis potently triggers complement activation. The purpose of this review is to summarise recent findings on complement activation and acquired brain or spinal cord injury, i.e. ischaemic-reperfusion injury or stroke, traumatic brain injury (TBI) and spinal cord injury (SCI), highlighting the potential for complement-targeted therapeutics to alleviate the devastating consequences of these neurological conditions

Impact of the Environment upon the Candida albicans Cell Wall and Resultant Effects upon Immune Surveillance

Acknowledgements This work was funded by a programme grant from the UK Medical Research Council [www.mrc.ac.uk: MR/M026663/1], and by PhD studentships from the University of Aberdeen to AP, DL. The work was also supported by the Medical Research Council Centre for Medical Mycology (MR/N006364/1) and by the Wellcome Trust [www.wellcome.ac.uk: 097377]. The funders had no role in study design, data collection and analysis, decision to publish or preparation of the manuscript.Postprin