Busting biofilms: free-living amoebae disrupt preformed methicillin-resistant Staphylococcus aureus (MRSA) and Mycobacterium bovis biofilms.

Biofilm-associated infections are difficult to eradicate because of their ability to tolerate antibiotics and evade host immune responses. Amoebae and/or their secreted products may provide alternative strategies to inhibit and disperse biofilms on biotic and abiotic surfaces. We evaluated the potential of five predatory amoebae - Acanthamoeba castellanii, Acanthamoeba lenticulata, Acanthamoeba polyphaga, Vermamoeba vermiformis and Dictyostelium discoideum - and their cell-free secretions to disrupt biofilms formed by methicillin-resistant Staphylococcus aureus (MRSA) and Mycobacterium bovis. The biofilm biomass produced by MRSA and M. bovis was significantly reduced when co-incubated with A. castellanii, A. lenticulata and A. polyphaga, and their corresponding cell-free supernatants (CFS). Acanthamoeba spp. generally produced CFS that mediated biofilm dispersal rather than directly killing the bacteria; however, A. polyphaga CFS demonstrated active killing of MRSA planktonic cells when the bacteria were present at low concentrations. The active component(s) of the A. polyphaga CFS is resistant to freezing, but can be inactivated to differing degrees by mechanical disruption and exposure to heat. D. discoideum and its CFS also reduced preformed M. bovis biofilms, whereas V. vermiformis only decreased M. bovis biofilm biomass when amoebae were added. These results highlight the potential of using select amoebae species or their CFS to disrupt preformed bacterial biofilms.

Amoeboid swimming in a compliant channel.

Several prokaryotes and eukaryotic cells swim in the presence of deformable and rigid surfaces that form confinement. The most commonly observed examples from biological systems are motility of leukocytes and pathogens present within the blood suspension through a microvascular network, and locomotion of eukaryotic cells such as immune system cells and cancerous cells through interstices between soft interstitial cells and the extracellular matrix within the interstitial tissue. This motivated us to investigate numerically the flow dynamics of amoeboid swimming in a flexible channel. The effects of wall stiffness and channel confinement on the flow dynamics and swimmer motion are studied. The swimmer motion through the flexible channel is substantially decelerated compared to the rigid channel. The strong confinement in the amply flexible channel imprisons the swimmer by severely restricting its forward motion. The swimmer velocity in a stiff channel displays nonmonotonic variation with the confinement while it shows monotonic reduction in a highly flexible channel. The physical rationale behind such distinct velocity behaviour in flexible and rigid channels is illustrated using an instantaneous flow field and flow history displayed by the swimmer. This behavior follows from a subtle interplay between the shape changes exhibited by the swimmer and the wall compliance. This study may aid in understanding the influence of elasticity of the surrounding environment on cell motility in immunological surveillance and invasiveness of cancer cells.

Transcriptome analyses of amoebic gill disease-affected Atlantic salmon (Salmo salar) tissues reveal localized host gene suppression.

The transcriptome response of Atlantic salmon (Salmo salar) displaying advanced stages of amoebic gill disease (AGD) was investigated. Naïve smolt were challenged with AGD for 19 days, at which time all fish were euthanized and their severity of infection quantified through histopathological scoring. Gene expression profiles were compared between heavily infected and naïve individuals using a 17 K Atlantic salmon cDNA microarray with real-time quantitative RT-PCR (qPCR) verification. Expression profiles were examined in the gill, anterior kidney, and liver. Twenty-seven transcripts were significantly differentially expressed within the gill; 20 of these transcripts were down-regulated in the AGD-affected individuals compared with naïve individuals. In contrast, only nine transcripts were significantly differentially expressed within the anterior kidney and five within the liver. Again the majority of these transcripts were down-regulated within the diseased individuals. A down-regulation of transcripts involved in apoptosis (procathepsin L, cathepsin H precursor, and cystatin B) was observed in AGD-affected Atlantic salmon. Four transcripts encoding genes with antioxidant properties also were down-regulated in AGD-affected gill tissue according to qPCR analysis. The most up-regulated transcript within the gill was an unknown expressed sequence tag (EST) whose expression was 218-fold (+/- SE 66) higher within the AGD affected gill tissue. Our results suggest that Atlantic salmon experiencing advanced stages of AGD demonstrate general down-regulation of gene expression, which is most pronounced within the gill. We propose that this general gene suppression is parasite-mediated, thus allowing the parasite to withstand or ameliorate the host response.

Monopylocystis visvesvarai n. gen., n. sp. and Sawyeria marylandensis n. gen., n. sp.: two new amitochondrial heterolobosean amoebae from anoxic environments.

Two new species of heterolobosean amoebae from anoxic environments, Monopylocystis visvesvarai and Sawyeria marylandensis, are described on the basis of light microscopy, electron microscopy, and their phylogenetic affiliation based on analyses of nuclear small-subunit ribosomal RNA gene sequences. Both species lack mitochondria but have organelles provisionally interpreted as hydrogenosomes, and neither can tolerate aerobic conditions. As their conditions of culture do not exclude all oxygen, they may be microaerophiles rather than strict anaerobes. Both species have unusual nucleolar morphologies. Monopylocystis visvesvarai, from a marine sediment, has nucleolar material distributed around the nuclear periphery. It is the first non-aerobic heterolobosean protist for which a cyst is known; the cyst is unmineralized and unornamented except for a single, raised, plugged pore. Sawyeria marylandensis, from an iron-rich freshwater stream, has nucleolar material distributed in one or two parietal masses, which persist during mitosis. In phylogenetic analyses of small-subunit rRNA gene sequences, Monopylocystis visvesvarai, Sawyeria marylandensis and Psalteriomonas lanterna converge to form a single clade of non-aerobic (anaerobic/microaerophilic) heteroloboseans.

The origins of parasitism in the protists.

The origins of parasitism among the protists are, like the group itself, polyphyletic. Probably the majority of present-day parasitic forms evolved from free-living ancestors which were ingested as part of the food of their hosts, though origins from ectoparasitic forms and via a phase of facultative parasitism are possibilities, particularly among the ciliated protozoa and (for ectoparasitism) the Kinetoplasta. Sporozoan parasites most probably developed via a stage which was ingested and became adapted to life in the host's gut. Further developments in parasitism involved deeper penetration into the host's tissues and the adoption of more than one host in the life cycle, thus avoiding entirely the potentially hazardous phase of existence outside the host.