Project description:Legionella pneumophila are important opportunistic pathogens for which environmental reservoirs such as protists are crucial for the infection of humans. Free-living amoebae are considered key hosts providing nutrients and shelter for highly efficient intracellular proliferation of L. pneumophila, which eventually leads to lysis of the amoeba host cell. Yet, the significance of other bacterial players for L. pneumophila ecology is poorly understood. In this study we used a ubiquitous amoeba and their bacterial endosymbiont to investigate the impact of this common association on L. pneumophila infection. We demonstrate that Acanthamoeba castellanii harboring the chlamydial symbiont Protochlamydia amoebophila were able to erase L. pneumophila and, in contrast to symbiont-free amoebae, survived the infection and were able to resume growth. Environmental amoeba isolates harboring P. amoebophila were equally well-protected, and fresh environmental isolates of L. pneumophila were equally well-erased, suggesting ecological relevance of this symbiont-mediated protection. We further show that protection was not mediated by impaired L. pneumophila uptake. Instead, we observed reduced virulence of L. pneumophila released from symbiont-containing amoebae that is strongly supported by transcriptome data. Interference with transition to the transmissive phase is thus likely the basis for this protection. Finally, our data indicate that the defensive response of amoebae harboring P. amoebophila leaves the amoebae with superior fitness reminiscent of immunological memory. Given that mutualistic associations between bacteria and amoebae are widely distributed, P. amoebophila and potentially other amoeba endosymbionts could be key elements in shaping environmental survival, abundance and virulence of this important pathogen thereby affecting frequency of human infection.
Project description:This experiment exploits the life-cycle of Strongyloides ratti, which is a parasitic nematode of brown rats that exhibits three adult stages within its life-cycle - parasitic females, freeliving females and free-living males. We use a cDNA microarray to examine patterns of (i) gender-biased gene expression by contrasting free-living females against free-living males, and (ii) parasitic-biased expression by contrasting parasitic females against free-living females. Of the 3688 distinct transcripts represented on our array, 20% exhibited male-biased expression 19% exhibit female-biased expression, 11% exhibit parasitic-biased expression and 8% exhibit free-living-biased expression. Among the top responding genes, an orthologue of major sperm protein is upregulated in males, distinct aspartic protease orthologues are upregulated in either parasitic or in free-living females, and orthologues of hsp-17 chaperone are upregulated in parasitic females. Upon a global analysis of gene expression, we find that female-biased expression is associated with genes involved in reproductive processes and larval development, that male-biased expression is associated with genes involved in metabolism, and that free-living biased expression is associated with genes involved in regulation of body fluids and response to external stimulus. The association of gene ontology with parasite-biased expression is less clear. Our results provide an initial gene expression analysis of gender- and parasite-biased expression in S. ratti, may be more generally applicable to other parasitic nematodes, and may help to refine the search for novel drug or vaccine targets against parasitic nematodes.
Project description:Here, we sought to test the resistance of human pathogens to unaltered environmental free-living amoebae. Amoebae are ubiquitous eukaryotic microorganisms and important predators of bacteria. Environmental amoebae have also been proposed to serve as both potential reservoirs and training grounds for human pathogens. However, studies addressing their relationships with human pathogens often rely on a few domesticated amoebae that have been selected to feed on rich medium, thereby possibly overestimating the resistance of pathogens to these predatory phagocytes. From an open-air composting site, we recovered over 100 diverse amoebae that were able to feed on Acinetobacter baumannii and Klebsiella pneumoniae. In a standardized and quantitative assay for predation, the isolated amoebae showed a broad predation spectrum, killing clinical isolates of A. baumannii, K. pneumoniae, Pseudomonas aeruginosa, and Staphylococcus aureus. Interestingly, A. baumannii, which was previously reported to resist predation by laboratory strains of Acanthamoeba, was efficiently consumed by closely related environmental amoebae. The isolated amoebae were capable of feeding on highly virulent carbapenem-resistant or methicillin-resistant clinical isolates. In conclusion, the natural environment is a rich source of amoebae with broad-spectrum bactericidal activities, including against antibiotic-resistant isolates. IMPORTANCE Free-living amoebae have been proposed to play an important role in hosting and disseminating various human pathogens. The resistance of human pathogens to predation by amoebae is often derived from in vitro experiments using model amoebae. Here, we sought to isolate environmental amoebae and to test their predation on diverse human pathogens, with results that challenge conclusions based on model amoebae. We found that the natural environment is a rich source of diverse amoebae with broad-spectrum predatory activities against human pathogens, including highly virulent and antibiotic-resistant clinical isolates.
Project description:In previous studies we have shown that the two adult females morphs of S. ratti have very different lifespans. This experiment was designed to try to identify differentially expressed genes in these two adult morphs that may account for these differing lifespans. The genes expressed by S. ratti parasitic females at day 6 p.i. were compared to the genes expressed by S. ratti free living females at 3 days 19 degrees C. This comparison was done using a microarray chip that is spotted with PCR fragments from the libraries that were generated from parasitic females extracted at day 6 and day 15 p.i., and a microarray chip that is spotted with PCR fragments from the libraries that were generated from free-living larval stages L1, L2 and infective L3s and from free-living males and females.
Project description:Mesorhizobium huakuii 7653R is an M-NM-1-proteobacterium that occurs either in a nitrogen-fixing symbiosis with its host plant, A. sinicus, or free-living in the soil. Investigation of whole genome gene expression level changes in Bacteroids compared to the free-living cells. Understand how M. huakuii 7653R responds to alterations in its environment and to the physiological changes that occur during bacteroid differentiation. Examination of mRNA levels in free-living cells and bacteroids at 32 days postinoculation
Project description:Free-living bacteria were grown on succinae ammonia AMS and gene expression was compared to free-living bacteria grown on glucose ammonia AMS.
Project description:Mesorhizobium huakuii 7653R is an α-proteobacterium that occurs either in a nitrogen-fixing symbiosis with its host plant, A. sinicus, or free-living in the soil. Investigation of whole genome gene expression level changes in Bacteroids compared to the free-living cells. Understand how M. huakuii 7653R responds to alterations in its environment and to the physiological changes that occur during bacteroid differentiation.