The development and characterization of an E. coli O25B bioconjugate vaccine

Extraintestinal pathogenic Escherichia coli (ExPEC) cause a wide range of clinical diseases such as bacteremia and urinary tract infections. The increase of multidrug resistant ExPEC strains is becoming a major concern for the treatment of these infections and E. coli has been identified as a critical priority pathogen by the WHO. Therefore, the development of vaccines has become increasingly important, with the surface lipopolysaccharide constituting a promising vaccine target. This study presents genetic and structural analysis of clinical urine isolates from Switzerland belonging to the serotype O25. Approximately 75% of these isolates were shown to correspond to the substructure O25B only recently described in an emerging clone of E. coli sequence type 131. To address the high occurrence of O25B in clinical isolates, an O25B glycoconjugate vaccine was prepared using an E. coli glycosylation system. The O antigen cluster was integrated into the genome of E. coli W3110, thereby generating an E. coli strain able to synthesize the O25B polysaccharide on a carrier lipid. The polysaccharide was enzymatically conjugated to specific asparagine side chains of the carrier protein exotoxin A (EPA) of Pseudomonas aeruginosa by the PglB oligosaccharyltransferase from Campylobacter jejuni. Detailed characterization of the O25B-EPA conjugate by use of physicochemical methods including NMR and GC-MS confirmed the O25B polysaccharide structure in the conjugate, opening up the possibility to develop a multivalent E. coli conjugate vaccine containing O25B-EPA.

     

CD4+CD25+ regulatory T cells in HIV infection

The immune system faces the difficult task of discerning between foreign, potentially pathogen-derived antigens and self-antigens. Several mechanisms, including deletion of self-reactive T cells in the thymus, have been shown to contribute to the acceptance of self-antigens and the reciprocal reactivity to foreign antigens. Over the last decade it has become increasingly clear that CD4(+)CD25(+) T(Reg) cells are crucial for maintenance of T cell tolerance to self-antigens in the periphery, and to avoid development of autoimmune disorders. Recently, evidence has also emerged that demonstrates that CD4(+)CD25(+) T(Reg) cells can also suppress T cell responses to foreign pathogens, including viruses such as HIV. In this article we review the current knowledge and potential role of CD4(+)CD25(+) T(Reg) cells in HIV infection.     

Assessment of self-organizing maps to analyze sole-carbon source utilization profiles

The use of community-level physiological profiles obtained with Biolog microplates is widely employed to consider the functional diversity of bacterial communities. Biolog produces a great amount of data which analysis has been the subject of many studies. In most cases, after some transformations, these data were investigated with classical multivariate analyses. Here we provided an alternative to this method, that is the use of an artificial intelligence technique, the Self-Organizing Maps (SOM, unsupervised neural network). We used data from a microcosm study of algae-associated bacterial communities placed in various nutritive conditions. Analyses were carried out on the net absorbances at two incubation times for each substrates and on the chemical guild categorization of the total bacterial activity. Compared to Principal Components Analysis and cluster analysis, SOM appeared as a valuable tool for community classification, and to establish clear relationships between clusters of bacterial communities and sole-carbon sources utilization. Specifically, SOM offered a clear bidimensional projection of a relatively large volume of data and were easier to interpret than plots commonly obtained with multivariate analyses. They would be recommended to pattern the temporal evolution of communities' functional diversity.     

Lrp-DNA complex stability determines the level of ON cells in type P fimbriae phase variation

F165(1) and the pyelonephritis-associated pili (Pap) are two members of the type P family of adhesive factors that play a key role in the establishment of disease caused by extraintestinal Escherichia coli (ExPEC) strains. They are both under the control of an epigenetic and reversible switch that defines the number of fimbriated (ON) and afimbriated (OFF) cells within a clonal population. Our present study demonstrates that the high level of ON cells found during F165(1) phase variation is due to altered stability of the DNA complex formed by the leucine-responsive regulatory protein (Lrp) at its repressor binding sites 1-3; after each cell cycle, complex formation is also modulated by the local regulator FooI (homologue to PapI) which promotes the transit of Lrp towards its activator binding sites 4-6. Furthermore, we identified two nucleotides (T490, G508) surrounding the Lrp binding site 1 that are critical to maintaining a high OFF to ON switch rate during F165(1) phase variation, as well as switching Pap fimbriae towards the OFF state.     

Unregulated hunting and genetic recovery from a severe population decline: the cautionary case of Bulgarian wolves

European wolf (Canis lupus) populations have suffered extensive decline and range contraction due to anthropogenic culling. In Bulgaria, although wolves are still recovering from a severe demographic bottleneck in the 1970s, hunting is allowed with few constraints. A recent increase in hunting pressure has raised concerns regarding long-term viability. We thus carried out a comprehensive conservation genetic analysis using microsatellite and mtDNA markers. Our results showed high heterozygosity levels (0.654, SE 0.031) and weak genetic bottleneck signals, suggesting good recovery since the 1970s decline. However, we found high levels of inbreeding (F _IS  = 0.113, SE 0.019) and a N _e/N ratio lower than expected for an undisturbed wolf population (0.11, 95 % CI 0.08–0.29). We also found evidence for hybridisation and introgression from feral dogs (C. familiaris) in 10 out of 92 wolves (9.8 %). Our results also suggest admixture between wolves and local populations of golden jackals (C. aureus), but less extensive as compared with the admixture with dogs. We detected local population structure that may be explained by fragmentation patterns during the 1970s decline and differences in local ecological characteristics, with more extensive sampling needed to assess further population substructure. We conclude that high levels of inbreeding and hybridisation with other canid species, which likely result from unregulated hunting, may compromise long-term viability of this population despite its current high genetic diversity. The existence of population subdivision warrants an assessment of whether separate management units are needed for different subpopulations. Our study highlights conservation threats for populations with growing numbers but subject to unregulated hunting.     

New fossils of ascomycetous anamorphic fungi from Baltic amber

Three new fossils of saprotrophic fungi are presented and described from Baltic amber, dated to Eocene epoch (Paleogene, upper to mid-Eocene). All belong to Ascomycota and are represented by hyphae as well as asexual reproduction structures allowing to assign them to present genera, respectively Periconia, Penicillium and Scopulariopsis. These material provide both the first and the oldest known fossil record of the mentioned taxa, making these data valuable for the knowledge about the evolutionary history of the Ascomycota.     

Quantitative determination of creatine kinase release from herring (Clupea harengus) spermatozoa induced by tributyltin

Creatine kinase (CK, ATP creatine phosphotransferase, EC 2.7.3.2) is an enzyme participating in ATP regeneration, which is the primary source of energy in living organisms. We demonstrated that CK from herring spermatozoa has high activity ( approximately 452 micromol/min/g of fresh semen) and has a different electrophoretic mobility from isoenzymes present in skeletal muscle. In our study, we investigated toxic effect of tributyltin (TBT) on herring spermatozoa using a specific sperm viability kit to observe live and dead sperm cells with a confocal microscope. Treatment of herring spermatozoa with TBT caused a time-dependent decrease of viability: 35% nonviable cells with 5 microM TBT and more than 90% nonviable cells with 10 microM TBT after 6 h exposure. We also monitored CK release from damaged spermatozoa into surrounding medium containing different concentrations of TBT. The higher concentration of TBT was used the more CK release from spermatozoa was observed. We suggest that CK could be a good biomarker of sperm cell membranes degradation in the case when lactate dehydrogenase release from permeabilized cells is not possible for rapid determination of the effect of TBT.