Testate amoebae as indicators of hydroseral change: an 8,500 year record from Mer Bleue Bog, eastern Ontario, Canada.

Testate amoebae have been used widely as a proxy of hydrological change in ombrotrophic peatlands, although their response to abiotic controls in other types of mire and fenland palaeo-environments is less well understood. This paper examines the response of testate amoebae to hydroseral and other environmental changes at Mer Bleue Bog, Ontario, Canada, a large ombrotrophic peatland, which evolved from a brackish-water embayment in the early Holocene. Sediments, plant macrofossils and diatoms examined from a 5.99 m core collected from the dome of the bog record six stages of development: i) a quiet, brackish-water riverine phase (prior to ca. 8500 cal BP); ii) a shallow lake (ca. 8500–8200 cal BP); iii) fen (8200–7600 cal BP); iv) transitional mire (7600–6900 cal BP); v) pioneer raised mire (6900–4450 cal BP); and vi) ombrotrophic bog (4450 cal BP-present).

Testate amoebae, notably small (<25 µm diameter) specimens of Centropyxis aculeata type, first appear in low abundances in sediments ascribed to the lacustrine phase. Diatoms from the same horizons record a shallowing in water depth, a decline in salinity and the development of emergent macrophytic vegetation, which may have provided favourable conditions for testate amoeba colonization. The testate amoeba communities of the inferred fen phase are more diverse and include centropyxids, cyclopyxids, Arcellidae and Hyalospheniidae, although the assemblages show some differences to those recently reported in modern European fen environments. The Fen–Bog Transition (FBT) is also dominated by C. aculeata type. The change in testate amoeba communities around this key transition is apparent in the results of Detrended Correspondence Analysis (DCA), and appears to reflect a latent nutrient gradient and a secondary moisture gradient. DCA analyses of plant macrofossil remains around the FBT show a similar trend, although the sensitivity of the two proxies to the inferred environmental changes differs. Comparisons with other regional mid-Holocene peatland records confirm the important influence of reduced effective precipitation on the testate amoeba communities during the initiation and development of Sphagnum-dominated, raised bog communities.

Mid to late Holocene hydrological change in continental eastern Canada: assessing regional trends from ombrotrophic peat records.

ABSTRACT High resolution records of mid-late Holocene hydro-climatic change are presented from Mer Bleue Bog, eastern Ontario. Past climatic changes in this region have previously been inferred from lake sediments, but rain-fed peatlands can offer additional insights into the spatial and temporal pattern of moisture availability. In this study, reconstructed water table depths are based on a testate amoeba-derived transfer function developed for the region and changes in bog surface wetness are compared with plant macrofossil and peat humification data.

RÉSUMÉ Nous présentons les enregistrements hautes résolutions des variations hydrologique durant la second moitié de l’Holocène pour les tourbières Mer Bleue á l’est de l'Ontario. Précédemment, les changements climatiques de cette région ont été dérivés à partir de prélèvement de sédiments de lac. Mais ils s’avèrent que les tourbières ombrotrophes offrir un éclairage supplémentaire sur les schémas de répartition spatiale et temporelle de la disponibilité de l'humidité. Dans cette étude, des profondeurs reconstruites de nappe phréatique sont basées sur un modèle de function de transfert d’amibes (Arcellinida) et des changements de l’humidité de surface de la tourbière sont comparés avec les macrofossils et au humification de tourbe dans une analyse multi-proxy.

A novel sensor kinase-response regulator hybrid controls biofilm formation and type VI secretion system activity in Burkholderia cenocepacia

Burkholderia cenocepacia is an important opportunistic pathogen causing serious chronic infections in patients with cystic fibrosis (CF). Adaptation of B. cenocepacia to the CF airways may play an important role in the persistence of the infection. We have identified a sensor kinase-response regulator (BCAM0379) named AtsR in B. cenocepacia K56-2 that shares 19% amino acid identity with RetS from Pseudomonas aeruginosa. atsR inactivation led to increased biofilm production and a hyperadherent phenotype in both abiotic surfaces and lung epithelial cells. Also, the atsR mutant overexpressed and hypersecreted an Hcp-like protein known to be specifically secreted by the type VI secretion system (T6SS) in other gram-negative bacteria. Amoeba plaque assays demonstrated that the atsR mutant was more resistant to Dictyostelium predation than the wild-type strain and that this phenomenon was T6SS dependent. Macrophage infection assays also demonstrated that the atsR mutant induces the formation of actin-mediated protrusions from macrophages that require a functional Hcp-like protein, suggesting that the T6SS is involved in actin rearrangements. Three B. cenocepacia transposon mutants that were found in a previous study to be impaired for survival in chronic lung infection model were mapped to the T6SS gene cluster, indicating that the T6SS is required for infection in vivo. Together, our data show that AtsR is involved in the regulation of genes required for virulence in B. cenocepacia K56-2, including genes encoding a T6SS.

Intracellular survival of Burkholderia cenocepacia in macrophages is associated with a delay in the maturation of bacteria-containing vacuoles

Strains of the Burkholderia cepacia complex (Bcc) are opportunistic bacteria that can cause life-threatening infections in patients with cystic fibrosis and chronic granulomatous disease. Previous work has shown that Bcc isolates can persist in membrane-bound vacuoles within amoeba and macrophages without bacterial replication, but the detailed mechanism of bacterial persistence is unknown. In this study, we have investigated the survival of the Burkholderia cenocepacia strain J2315 within RAW264.7 murine macrophages. Strain J2315 is a prototypic isolate of the widespread and transmissible ET12 clone. Unlike heat-inactivated bacteria, which reach lysosomes shortly after internalization, vacuoles containing live B. cenocepacia J2315 accumulate the late endosome/lysosome marker LAMP-1 and start fusing with lysosomal compartments only after 6 h post internalization. Using fluorescent fluid-phase probes, we also demonstrated that B. cenocepacia-containing vacuoles continued to interact with newly formed endosomes, and maintained a luminal pH of 6.4 +/- 0.12. In contrast, vacuoles containing heat-inactivated bacteria had an average pH of 4.8 +/- 0.03 and rapidly merged with lysosomes. Additional experiments using concanamycin A, a specific inhibitor of the vacuolar H+-ATPase, revealed that vacuoles containing live bacteria did not exclude the H+-ATPase. This mode of bacterial survival did not require type III secretion, as no differences were found between wild type and a type III secretion mutant strain. Collectively, our results suggest that intracellular B. cenocepacia cause a delay in the maturation of the phagosome, which may contribute to facilitate bacterial escape from the microbicidal activities of the host cell.

Role of Bacterial Surface Structures on the Interaction of Klebsiella pneumoniae with Phagocytes

Phagocytosis is a key process of the immune system. The human pathogen Klebsiella pneumoniae is a well known example of a pathogen highly resistant to phagocytosis. A wealth of evidence demonstrates that the capsule polysaccharide (CPS) plays a crucial role in resistance to phagocytosis. The amoeba Dictyostelium discoideum shares with mammalian macrophages the ability to phagocytose and kill bacteria. The fact that K. pneumoniae is ubiquitous in nature and, therefore, should avoid predation by amoebae, poses the question whether K. pneumoniae employs similar means to counteract amoebae and mammalian phagocytes. Here we developed an assay to evaluate K. pneumoniae-D. discoideum interaction. The richness of the growth medium affected the threshold at which the cps mutant was permissive for Dictyostelium and only at lower nutrient concentrations the cps mutant was susceptible to predation by amoebae. Given the critical role of bacterial surface elements on host-pathogen interactions, we explored the possible contribution of the lipopolysaccharide (LPS) and outer membrane proteins (OMPs) to combat phagoyctosis by D. discoideum. We uncover that, in addition to the CPS, the LPS O-polysaccharide and the first core sugar participate in Klebsiella resistance to predation by D. discoideum. K. pneumoniae LPS lipid A decorations are also necessary to avoid predation by amoebae although PagP-dependent palmitoylation plays a more important role than the lipid A modification with aminoarabinose. Mutants lacking OMPs OmpA or OmpK36 were also permissive for D. discoideium growth. Except the LPS O-polysaccharide mutants, all mutants were more susceptible to phagocytosis by mouse alveolar macrophages. Finally, we found a correlation between virulence, using the pneumonia mouse model, and resistance to phagocytosis. Altogether, this work reveals novel K. pneumoniae determinants involved in resistance to phagocytosis and supports the notion that Dictyostelium amoebae might be useful as host model to measure K. pneumoniae virulence and not only phagocytosis. © 2013 March et al.

Evaluating the use of testate amoebae for palaeohydrological reconstruction in permafrost peatlands

The melting of high-latitude permafrost peatlands is a major concern due to a potential positive feedback on global climate change. We examine the ecology of testate amoebae in permafrost peatlands, based on sites in Sweden (~ 200 km north of the Arctic Circle). Multivariate statistical analysis confirms that water-table depth and moisture content are the dominant controls on the distribution of testate amoebae, corroborating the results from studies in mid-latitude peatlands. We present a new testate amoeba-based water table transfer function and thoroughly test it for the effects of spatial autocorrelation, clustered sampling design and uneven sampling gradients. We find that the transfer function has good predictive power; the best-performing model is based on tolerance-downweighted weighted averaging with inverse deshrinking (performance statistics with leave-one-out cross validation: R2 = 0.87, RMSEP = 5.25 cm). The new transfer function was applied to a short core from Stordalen mire, and reveals a major shift in peatland ecohydrology coincident with the onset of the Little Ice Age (c. AD 1400). We also applied the model to an independent contemporary dataset from Stordalen and find that it outperforms predictions based on other published transfer functions. The new transfer function will enable palaeohydrological reconstruction from permafrost peatlands in Northern Europe, thereby permitting greatly improved understanding of the long-term ecohydrological dynamics of these important carbon stores as well as their responses to recent climate change.