PdhS,an Old-Pole-Localized Histidine Kinase,Recruits the Fumarase FumC in Brucella abortus

The bacterial pathogen Brucella abortus was recently demonstrated to recruit the essential cytoplasmic histidine kinase PdhS to its old pole. Here, we report identification of the fumarase FumC as a specific partner for the N-terminal “sensing” domain of PdhS, using an ORFeome-based yeast two-hybrid screen. We observed that FumC and PdhS colocalize at the old pole of B. abortus, while the other fumarase FumA is not polarly localized. FumC is not required for PdhS localization, and polar FumC localization is not FumA dependent. FumC homologs are not polarly localized in Sinorhizobium meliloti and Caulobacter crescentus, suggesting that polar recruitment of FumC by PdhS is evolutionarily recent.It was previously reported that morphological asymmetry, here defined by the production of two sibling cells different in length at each cell cycle, is a common feature shared by at least four members of the alphaproteobacteria with diversified lifestyles, namely, the free-living model bacterium Caulobacter crescentus, the plant pathogen Agrobacterium tumefaciens, the plant symbiont Sinorhizobium meliloti, and the facultative intracellular pathogen Brucella abortus (5). In B. abortus, an essential cytoplasmic histidine kinase named PdhS is anchored to the old pole of the bacterium (6). PdhS interacts with the response regulator DivK (6).PdhS is a rather atypical histidine kinase. It is cytoplasmic and contains an N-terminal “sensing” domain with no predicted function, except for a PAS domain that remains to be characterized. Nevertheless, the N-terminal part preceding the PAS domain of PdhS is sufficient for polar localization in B. abortus, as well as in the alphaproteobacteria S. meliloti and C. crescentus (6). This suggests that a molecular mechanism and/or polar structure required for PdhS targeting is conserved in these alphaproteobacteria.     

Sublethal injuries in Middle Jurassic ammonite shells from Poland

Sublethal injuries, the effects of which are seen as regeneration patterns, are described from Late Bajocian and Bathonian (Middle Jurassic) ammonites from Poland (Polish Jura area) for the first time. The total number of ammonite shells bearing signs of sublethal injuries is small (only 11 specimens, which constitute ∼1.2% of all ammonites investigated), and this value is even smaller (∼0.3 to 0.8%) when analysing a large sample of a particular ammonite species. Specimens under consideration represent ten species, belonging to six genera and five families. All the healed injuries are represented by only one type, referred to as the ‘forma verticata’ of Hölder. This type of regeneration, very common in ammonite shells in general, is an effect of a puncture injury of the shell-secreting mantle-epithelia at the apertural margin. Although many different extrinsic (mechanical) factors may be responsible for such healed injuries, here it is most plausible they are an effect of either competitive or predatory activities. Other causes, like collision of the ammonite shells with the substrate in a high-energy environment, are excluded because the sea-bottom was soft and situated below the storm wave-base. From many potential predators inhabiting the Polish Basin during the Bajocian and Bathonian, the most likely to have caused these injuries are other ammonites, belemnites and nautiloids. Crabs, which are cited in the literature as a probable perpetrator of the ‘forma verticata’ injuries, appear unlikely here, as the ammonites under discussion were not purely benthic.     

Pathophysiological Mechanisms of Severe Anaemia in Malawian Children

Background

Severe anaemia is a major cause of morbidity and mortality in African children. The aetiology is multi-factorial, but interventions have often targeted only one or a few causal factors, with limited success.

Methods and Findings

We assessed the contribution of different pathophysiological mechanisms (red cell production failure [RCPF], haemolysis and blood loss) to severe anaemia in Malawian children in whom etiological factors have been described previously. More complex associations between etiological factors and the mechanisms were explored using structural equation modelling. In 235 children with severe anaemia (haemoglobin<3.2 mMol/L [5.0 g/dl]) studied, RCPF, haemolysis and blood loss were found in 48.1%, 21.7% and 6.9%, respectively. The RCPF figure increased to 86% when a less stringent definition of RCPF was applied. RCPF was the most common mechanism in each of the major etiological subgroups (39.7–59.7%). Multiple aetiologies were common in children with severe anaemia. In the final model, nutritional and infectious factors, including malaria, were directly or indirectly associated with RCPF, but not with haemolysis.

Conclusion

RCPF was the most common pathway leading to severe anaemia, from a variety of etiological factors, often found in combination. Unlike haemolysis or blood loss, RCPF is a defect that is likely to persist to a significant degree unless all of its contributing aetiologies are corrected. This provides a further explanation for the limited success of the single factor interventions that have commonly been applied to the prevention or treatment of severe anaemia. Our findings underline the need for a package of measures directed against all of the local aetiologies of this often fatal paediatric syndrome.     

Intracellular degradation of somatostatin-14 following somatostatin-receptor3-mediated endocytosis in rat insulinoma cells

Somatostatin receptor (SSTR) endocytosis influences cellular responsiveness to agonist stimulation and somatostatin receptor scintigraphy, a common diagnostic imaging technique. Recently, we have shown that SSTR1 is differentially regulated in the endocytic and recycling pathway of pancreatic cells after agonist stimulation. Additionally, SSTR1 accumulates and releases internalized somatostatin-14 (SST-14) as an intact and biologically active ligand. We also demonstrated that SSTR2A was sequestered into early endosomes, whereas internalized SST-14 was degraded by endosomal peptidases and not routed into lysosomal degradation. Here, we examined the fate of peptide agonists in rat insulinoma cells expressing SSTR3 by biochemical methods and confocal laser scanning microscopy. We found that [(125)I]Tyr11-SST-14 rapidly accumulated in intracellular vesicles, where it was degraded in an ammonium chloride-sensitive manner. In contrast, [(125)I]Tyr1-octreotide accumulated and was released as an intact peptide. Rhodamine-B-labeled SST-14, however, was rapidly internalized into endosome-like vesicles, and fluorescence signals colocalized with the lysosomal marker protein cathepsinD. Our data show that SST-14 was cointernalized with SSTR3, was uncoupled from the receptor, and was sorted into an endocytic degradation pathway, whereas octreotide was recycled as an intact peptide. Chronic stimulation of SSTR3 also induced time-dependent downregulation of the receptor. Thus, the intracellular processing of internalized SST-14 and the regulation of SSTR3 markedly differ from the events mediated by the other SSTR subtypes.     

Long term outcome of severe anaemia in Malawian children

Background

Severe anaemia is a common, frequently fatal, condition in African children admitted to hospital, but its long term outcome is unknown. Early reports that survivors may be at risk of additional late morbidity and mortality may have significant implications for child survival in Africa. We assessed the short and long term outcome of severe anaemia in Malawian children and identified potential risk factors for death and further severe anaemia.

Methodology and Findings

For 18 months, we followed up children (6–60 months old) presenting to hospital with severe anaemia (haemoglobin ≤5g/dl) and their hospital and community controls with the aim to compare all cause mortality and severe anaemia recurrence rates between the groups, and to identify risk factors for these adverse outcomes. A total of 377 cases, 377 hospital controls and 380 community controls were recruited. Among cases, the in-hospital mortality was 6.4% and post-discharge all cause mortality was 12.6%, which was significantly greater than in hospital controls (2.9%) or community controls (1.4%) (Log rank test, p<0.001). The incidence of recurrence of severe anaemia among the cases was 0.102 per child-year (95% Confidence Interval 0.075–0.138), and was significantly higher than the 0.007 per child-year (95% CI 0.003–0.015) in the combined controls (p<0.0001). HIV was the most important risk factor both for post-discharge mortality (Hazard Ratio 10.5, 95% CI 4.0–27.2) and for recurrence of severe anaemia (HR 5.6, 95% CI 1.6–20.1).

Conclusions

Severe anaemia carries a high ‘hidden’ morbidity and mortality occurring in the months after initial diagnosis and treatment. Because severe anaemia is very common, this is likely to contribute importantly to overall under-five mortality. If not adequately addressed, severe anaemia may be an obstacle to achievement of the Millennium development goal No.4 on child survival. Strategies to diagnose and properly treat HIV infected children early most likely will reduce the high post-discharge mortality in severe anaemia.     

Alkalispirillum mobile gen. nov., spec. nov., an alkaliphilic non-phototrophic member of the Ectothiorhodospiraceae

From cultures of the anoxygenic phototroph Halorhodospira halophila SL-1, an aerobic, gram-negative spirillum was isolated. This moderately halophilic, alkaliphilic bacterium was motile by means of a single polar flagellum. It is described here as Alkalispirillum mobile gen. nov., spec. nov. Phylogenetic analysis of the Alkalispirillum mobile 16S rRNA gene led to its classification in the gamma-subclass of the Proteobacteria, as it appears closely related to phototrophic purple sulfur bacteria of the genera Ectothiorhodospira and Halorhodospira. Surprisingly, A. mobile is an obligate aerobe. The organism grows optimally with a number of carboxylic acids (such as sodium acetate) as carbon source, at 2% (i.e. approximately 0.34 M) sodium chloride, at pH 9-10, and at temperatures ranging from 35 to 38 degrees C. The dominant cellular fatty acids of Alkalispirillum mobile are C12:0, C16:0, C18:1cis11, and C18:0; its G+C content is 66.2+/-0.5 mol%.     

Directly monitoring individual retrovirus budding events using atomic force microscopy

Retrovirus budding is a key step in the virus replication cycle. Nonetheless, very little is known about the underlying mechanism of budding, primarily due to technical limitations preventing visualization of bud formation in real time. Methods capable of monitoring budding dynamics suffer from insufficient resolution, whereas other methods, such as electron microscopy, do not have the ability to operate under physiological conditions. Here we applied atomic force microscopy to real-time visualization of individual Moloney murine leukemia virus budding events. By using a single-particle analysis approach, we were able to observe distinct patterns in budding that otherwise remain transparent. We find that bud formation follows at least two kinetically distinct pathways. The majority of virions (74%) are produced in a slow process (>45 min), and the remaining particles (26%) assemble via a fast process (<25 min). Interestingly, repetitive budding from the same site was seen to occur in only two locations. This finding challenges the hypothesis that viral budding occurs from distinct sites and suggests that budding is not restricted laterally. In this study, we established a method to monitor the fine dynamics of the virus budding process. Using this single-particle analysis to study mutated viruses will enable us to gain additional insight into the mechanisms of viral budding.     

Cyrtocrinids (Crinoidea) and associated stalked crinoids from the Lower/Middle Oxfordian (Upper Jurassic) shelfal deposits of southern Poland

Five cyrtocrinid crinoid taxa previously unknown from the epicratonic deposits of Poland, as well as associated millericrinids and isocrinids, are described. The studied materials were derived mainly from the Lower and Middle Oxfordian, but crinoids are also from uppermost Callovian and/or lowermost Oxfordian sediments of the Polish Jura Chain (southern Poland). The crinoids, preserved as more or less complete (e.g., basal circlets) cups, include Lonchocrinus dumortieri, Phyllocrinus belbekensis, Remisovicrinus polonicus, Remisovicrinus aff. polonicus, Tetracrinus moniliformis and Sclerocrinus sp. The occurrence of Remisovicrinus polonicus in the late Middle Oxfordian of the southern Poland is confirmed. Moreover, the present study extends the geographic range of all cyrtocrinid species studied and discusses their unusual environmental adaptations.