In vitro multiplication in liquid culture of Syngonium contaminated with Bacillus spp. and Rathayibacter tritici

Contaminated Syngonium clusters were multiplied in an air lift bioreactor in liquid medium containing sucrose with the medium being circulated through a sterilizing filter. After 30 days, the culture in filtered medium produced 19.5 shoot initials per gram fresh weight of inoculum compared to 8.7 shoot initials produced in unfiltered medium. Transfer to an elongation medium with 30 mg l^-1 Rifampicin produced shoots on 67% of the clusters, while transfer to elongation medium without Rifampicin poduced shoots on 40% of the clusters. Clusters grown for three subcultures in a reactor without medium filtration had lost their multiplication ability. Clusters grown for three subcultures in a reactor with filtration, however, continued to show a two-three fold increase in fresh weight and shoot production.Abbreviations MS Murashige and Skoog     

Direct and indirect effects of dragonfly (<Emphasis Type="Italic">Anax imperator</Emphasis>) nymphs on green toad (<Emphasis Type="Italic">Bufo viridis</Emphasis>) tadpoles

We conducted an artificial pond experiment to assess the direct and indirect effects of predation on Bufo viridis tadpoles. We ran three treatments: free Anax (unrestrained predatory dragonfly nymph Anax imperator), caged Anax (non-consumptive effects), and control (no Anax). Anax showed both strong consumptive and non-consumptive effects on Bufo tadpoles. Free Anax eliminated all of the tadpoles within six days. Tadpoles preferred the shady side of the ponds. Caged Anax caused tadpoles to increase their spatial preferences. Tadpoles avoided the center of the pond, and in the presence of the caged predator, they were found in the center even less. Tadpoles also showed a strong preference for crowding together, and in the presence of a caged Anax, they tended to crowd more. Moreover, Bufo metamorphosed earlier and at a larger size in the caged Anax ponds, possibly by providing extra food resources due to the extra organic matter excreted by the predators. Handling editor: K. Martens     

Parkin promotes degradation of the mitochondrial pro-apoptotic ARTS protein

Parkinson's disease (PD) is associated with excessive cell death causing selective loss of dopaminergic neurons. Dysfunction of the Ubiquitin Proteasome System (UPS) is associated with the pathophysiology of PD. Mutations in Parkin which impair its E3-ligase activity play a major role in the pathogenesis of inherited PD. ARTS (Sept4_i2) is a mitochondrial protein, which initiates caspase activation upstream of cytochrome c release in the mitochondrial apoptotic pathway. Here we show that Parkin serves as an E3-ubiquitin ligase to restrict the levels of ARTS through UPS-mediated degradation. Though Parkin binds equally to ARTS and Sept4_i1 (H5/PNUTL2), the non-apoptotic splice variant of Sept4, Parkin ubiquitinates and degrades only ARTS. Thus, the effect of Parkin on ARTS is specific and probably related to its pro-apoptotic function. High levels of ARTS are sufficient to promote apoptosis in cultured neuronal cells, and rat brains treated with 6-OHDA reveal high levels of ARTS. However, over-expression of Parkin can protect cells from ARTS-induced apoptosis. Furthermore, Parkin loss-of-function experiments reveal that reduction of Parkin causes increased levels of ARTS and apoptosis. We propose that in brain cells in which the E3-ligase activity of Parkin is compromised, ARTS levels increase and facilitate apoptosis. Thus, ARTS is a novel substrate of Parkin. These observations link Parkin directly to a pro-apoptotic protein and reveal a novel connection between Parkin, apoptosis, and PD.     

Real-time analysis of effector translocation by the type III secretion system of enteropathogenic Escherichia coli

Bacteria use type III secretion systems (TTSS) to translocate effector proteins into host cells. Better understanding of the TTSS and its effectors' functions will require assays to measure their activities in vivo and in real time. We designed a real-time, high-throughput translocation assay that utilizes fusions of effector genes to the beta-lactamase reporter gene, positioned under the effector's native promoter and chromosomal location. Using this assay, we simultaneously and quantitatively analyzed the translocation kinetics of six core enteropathogenic E. coli effectors, EspF, EspG, EspH, EspZ, Map, and Tir. A distinct order in the efficiencies of effector translocation was observed. Translocation efficiency was determined by multiple factors, including the intrabacterial effector concentration, effector-chaperone interactions, the efficiency of bacterial attachment to the host cells, and possibly also by a translocation autoinhibition mechanism. The described real-time translocation assay could be easily adapted for varied applications in the study of bacterial pathogenesis.     

Small RNAs encoded within genetic islands of Salmonella typhimurium show host-induced expression and role in virulence

The emergence of pathogenic strains of enteric bacteria and their adaptation to unique niches are associated with the acquisition of foreign DNA segments termed ‘genetic islands’. We explored these islands for the occurrence of small RNA (sRNA) encoding genes. Previous systematic screens for enteric bacteria sRNAs were mainly carried out using the laboratory strain Escherichia coli K12, leading to the discovery of ~80 new sRNA genes. These searches were based on conservation within closely related members of enteric bacteria and thus, sRNAs, unique to pathogenic strains were excluded. Here we describe the identification and characterization of 19 novel unique sRNA genes encoded within the ‘genetic islands’ of the virulent strain Salmonella typhimurium. We show that the expression of many of the island-encoded genes is associated with stress conditions and stationary phase. Several of these sRNA genes are induced when Salmonella resides within macrophages. One sRNA, IsrJ, was further examined and found to affect the translocation efficiency of virulence-associated effector proteins into nonphagocytic cells. In addition, we report that unlike the majority of the E. coli sRNAs that are trans regulators, many of the island-encoded sRNAs affect the expression of cis-encoded genes. Our study suggests that the island encoded sRNA genes play an important role within the network that regulates bacterial adaptation to environmental changes and stress conditions and thus controls virulence.