Salmonella biofilm development depends on the phosphorylation status of RcsB

The Rcs phosphorelay pathway is a complex signaling pathway involved in the regulation of many cell surface structures in enteric bacteria. In response to environmental stimuli, the sensor histidine kinase (RcsC) autophosphorylates and then transfers the phosphate through intermediary steps to the response regulator (RcsB), which, once phosphorylated, regulates gene expression. Here, we show that Salmonella biofilm development depends on the phosphorylation status of RcsB. Thus, unphosphorylated RcsB, hitherto assumed to be inactive, is essential to activate the expression of the biofilm matrix compounds. The prevention of RcsB phosphorylation either by the disruption of the phosphorelay at the RcsC or RcsD level or by the production of a nonphosphorylatable RcsB allele induces biofilm development. On the contrary, the phosphorylation of RcsB by the constitutive activation of the Rcs pathway inhibits biofilm development, an effect that can be counteracted by the introduction of a nonphosphorylatable RcsB allele. The inhibition of biofilm development by phosphorylated RcsB is due to the repression of CsgD expression, through a mechanism dependent on the accumulation of the small noncoding RNA RprA. Our results indicate that unphosphorylated RcsB plays an active role for integrating environmental signals and, more broadly, that RcsB phosphorylation acts as a key switch between planktonic and sessile life-styles in Salmonella enterica serovar Typhimurium.     

Redescriptions of two frequently recorded but poorly known hemiurid digeneans, Lecithochirium musculus (Looss, 1907) (Lecithochiriinae) and Ectenurus lepidus Looss, 1907 (Dinurinae), based on material from the western Mediterranean

Two frequently reported but poorly known hemiurid digeneans, Lecithochirium musculus (Looss, 1907) (Lecithochiriinae), from the stomach of Trachinus draco and Citharus linguatula, and Ectenurus lepidus Looss, 1907 (Dinurinae), from the stomach of Spicara maena, are redescribed based on material from off the Barcelona coast of the western Mediterranean. The two species are commented upon, and Lecithochirium israelense Fischthal, 1980 is considered a synonym of L. musculus. Records of the two species in the Mediterranean Basin and North East Atlantic region are summarised.     

The Differential Interaction of Brucella and Ochrobactrum with Innate Immunity Reveals Traits Related to the Evolution of Stealthy Pathogens

Background

During evolution, innate immunity has been tuned to recognize pathogen-associated molecular patterns. However, some α-Proteobacteria are stealthy intracellular pathogens not readily detected by this system. Brucella members follow this strategy and are highly virulent, but other Brucellaceae like Ochrobactrum are rhizosphere inhabitants and only opportunistic pathogens. To gain insight into the emergence of the stealthy strategy, we compared these two phylogenetically close but biologically divergent bacteria.

Methodology/Principal Findings

In contrast to Brucella abortus, Ochrobactrum anthropi did not replicate within professional and non-professional phagocytes and, whereas neutrophils had a limited action on B. abortus, they were essential to control O. anthropi infections. O. anthropi triggered proinflammatory responses markedly lower than Salmonella enterica but higher than B. abortus. In macrophages and dendritic cells, the corresponding lipopolysaccharides reproduced these grades of activation, and binding of O. anthropi lipopolysaccharide to the TLR4 co-receptor MD-2 and NF-κB induction laid between those of B. abortus and enteric bacteria lipopolysaccharides. These differences correlate with reported variations in lipopolysaccharide core sugars, sensitivity to bactericidal peptides and outer membrane permeability.

Conclusions/Significance

The results suggest that Brucellaceae ancestors carried molecules not readily recognized by innate immunity, so that non-drastic variations led to the emergence of stealthy intracellular parasites. They also suggest that some critical envelope properties, like selective permeability, are profoundly altered upon modification of pathogen-associated molecular patterns, and that this represents a further adaptation to the host. It is proposed that this adaptive trend is relevant in other intracellular α-Proteobacteria like Bartonella, Rickettsia, Anaplasma, Ehrlichia and Wolbachia.     

Effects of <Emphasis Type="Italic">Caulerpa racemosa</Emphasis> var. <Emphasis Type="Italic">cylindracea</Emphasis> on prey availability: an experimental approach to predation of amphipods by <Emphasis Type="Italic">Thalassoma pavo</Emphasis> (Labridae)

Alien plant species, such as Caulerpa racemosa var. cylindracea, that invade Mediterranean marine vegetated habitats can affect habitat structure. In turn, changes in habitat structure may affect the associated invertebrate assemblages, either through changes in habitat selection or as a result of altered predation efficiency. In order to test for effects of changes in habitat structure resulting from colonization by C. racemosa on prey availability for predators, the importance of amphipods as a trophic resource in natural vegetated habitat was first assessed, and later experiments were undertaken to assess the effects of the alien alga on predation by Thalassoma pavo of two dominant amphipods: Elasmopus brasiliensis (Gammaridea) and Caprella dilatata (Caprellidea). Laboratory experiments were conducted in separate aquaria with five vegetation habitat types: Halopteris scoparia, Jania rubens, C. racemosa without detritus, C. racemosa with detritus, Cymodocea nodosa, together with controls. The vegetation was first defaunated, and then 30 amphipods were introduced to each aquarium and exposed to a single Thalassoma pavo individual for 1 h, after which the fish’s gut contents were examined. Consumption (per fish per hour) of caprellids (11.7 ± 1.4) was higher overall than that of gammarids (8.7 ± 1.5) and likely reflects different microhabitat use by amphipods, which affects susceptibility to predators. Consumption of amphipods also varied by habitat type. The highest predation rate was found in the C. nodosa habitat (12.7 ± 2.19) and the lowest in the C. racemosa habitats with detritus (4.1 ± 1.78) and without detritus (5.2 ± 0.55), which did not differ. The pattern of predation across habitats, however, was similar for both caprellid and gammarid amphipods, indicating a more general effect of habitat on amphipod predation. Our findings showed that invasive species such as C. racemosa can decrease feeding by predators such as T. pavo. Changes in predator–prey interactions could have consequences for food web support in the Mediterranean.     

Histological survey of symbionts and other conditions in razor clam Ensis arcuatus (Jeffreys, 1865) (Pharidae) of the coast of Galicia (NW Spain)

Symbionts and abnormal conditions of razor clam Ensis arcuatus were surveyed in three commercially important natural beds of Galician estuaries (NW Spain). Samples of 15-20 E. arcuatus were collected every 2 months from January 2003 until July 2004 and processed for histological examination. Prokaryote-like colonies, renal coccidians, gregarines, Trichodina sp. ciliates, haplosporidian-like plasmodia, turbelaria, trematode metacercariae, cestode-like larvae and basophilic inclusion bodies were observed in razor clam tissues without causing host damage. Bucephalid digenean sporocysts and germinoma were seen in some samples causing moderate or severe damage to the host depending on the intensity of infection and both could be a cause for concern if prevalence reached epizootic levels in Galician E. arcuatus populations. None of the parasites detected is OIE notifiable and, in general, the commercially exploited beds studied seem to be devoid of serious pathogens.     

Sepsis induces DNA fragmentation in rat skeletal muscle

Recent studies have demonstrated the activation of skeletal muscle DNA fragmentation in some catabolic conditions. In an attempt to elucidate if sepsis (a catabolic state) was also associated with muscle apoptosis, sepsis was induced by cecal ligation and puncture, and the results clearly show an induction of DNA fragmentation in gastrocnemius muscle following the induction of the septic state. Administration of rolipram (an inhibitor of tumour necrosis factor-a (TNF-alpha) synthesis) to septic rats clearly prevented the increased DNA fragmentation, suggesting that TNF-alpha is involved in the activation of the apoptotic events in septic rat skeletal muscle.     

Heparin interferes with translocation of Yop proteins into HeLa cells and binds to LcrG, a regulatory component of the Yersinia Yop apparatus

Yersiniae are equipped with the Yop virulon, an apparatus that allows extracellular bacteria to deliver toxic Yop proteins inside the host cell cytosol in order to sabotage the communication networks of the host cell or even to cause cell death. LcrG is a component of the Yop virulon involved in the regulation of secretion of the Yops. In this paper, we show that LcrG can bind HeLa cells, and we analyse the role of proteoglycans in this phenomenon. Treatment of the HeLa cells with heparinase I, but not chondroitinase ABC, led to inhibition of binding. Competition assays indicated that heparin and dextran sulphate strongly inhibited binding, but that other glycosaminoglycans did not. This demonstrated that binding of HeLa cells to purified LcrG is caused by heparan sulphate proteoglycans. LcrG could bind directly to heparin-agarose beads and, in agreement with these results, analysis of the protein sequence of Yersinia enterocolitica LcrG revealed heparin-binding motifs. In vitro production and secretion by Y . enterocolitica of the Yops was unaffected by the addition of heparin. However, the addition of exogenous heparin decreased the level of YopE–Cya translocation into HeLa cells. A similar decrease was seen with dextran sulphate, whereas the other glycosaminoglycans tested had no significant effect. Translocation was also decreased by treatment of HeLa cells with heparinitase, but not with chondroitinase. Thus, heparan sulphate proteoglycans have an important role to play in translocation. The interaction between LcrG and heparan sulphate anchored at the surface of HeLa cells could be a signal triggering deployment of the Yop translocation machinery. This is the first report of a eukaryotic receptor interacting with the type III secretion and associated translocation machinery of Yersinia or of other bacteria.     

The Myf fibrillae of Yersinia enterocolitica

The Myf antigen produced by Yersinia enterocolitica appeared as a proteic polymer composed of 21 kDa subunits. By transposon mutagenesis we isolated Myf-defective mutants. Those allowed us to clone and sequence a 4.4 kb chromosomal locus involved in Myf production. This region was found to contain three genes that we called myfA, myfB and myfC. Genes myfB and myfC encode an assembly machine related to those involved in the synthesis of many fimbriae; MyfB, the putative chaperone, possesses the consensus residues of the PapD family and MyfC encodes a putative outer-membrane protein. MyfA, the major subunit, was found to be 44% identical to the pH 6 antigen of Y.pestis. Myf is thus the K enterocolitica counterpart of this antigen, but it is by far not so well conserved as the other virulence determinants such as the Yops, suggesting that Myf and pH 6 antigen do not necessarily play the same role in Y. enterocolitica and Y. pestis. The study of the prevalence of myfA in various species of Yersinia reveaied that, like the yst enterotoxin gene, its presence is restricted to the pathogenic serotypes of Y. enterocolitica. By immuno-gold labelling, Myf appeared as a layer of extracellular material extending locally 2μm from the bacterial surface, indicative of a fibrillar structure.     

Study of the interaction between a histidine-deoxyguanosine hybrid and cisplatin

 A histidine-2′-deoxyguanosine hybrid, Phac-Hse(p^5′dG)-His-OH (I), was synthesized, and its reaction with cisplatin was monitored by reversed-phase HPLC and ¹H NMR. Two new compounds, II and III, were observed to be simultaneously formed, II in larger proportion than III. These products were isolated after HPLC purification and extensively characterized. Both II and III contained platinum, had the same mass and showed a bathochromic displacement of their absorption maxima with respect to that of I. Both remained undegraded upon enzymatic digestion and yielded I when treated with NaCN. From these data and the information provided by ¹H NMR analysis, we inferred that II and III were constitutional isomers, in particular chelates in which platinum was coordinated to the N7 of guanine and either the N^π or the N^τ imidazole nitrogens, respectively. No preference of the metal for either of these N-donors was observed. Received: 3 May 1999 / Accepted: 24 August 1999