Type IV Pili in Francisella – A Virulence Trait in an Intracellular Pathogen

Francisella tularensis is a highly virulent intracellular human pathogen that is capable of rapid proliferation in the infected host. Mutants affected in intracellular survival and growth are highly attenuated which highlights the importance of the intracellular phase of the infection. Genomic analysis has revealed that Francisella encodes all genes required for expression of functional type IV pili (Tfp), and in this focused review we summarize recent findings regarding this system in the pathogenesis of tularemia. Tfp are dynamic adhesive structures that have been identified as major virulence determinants in several human pathogens, but it is not obvious what role these structures could have in an intracellular pathogen like Francisella. In the human pathogenic strains, genes required for secretion and assembly of Tfp and one pilin, PilA, have shown to be required for full virulence. Importantly, specific genetic differences have been identified between the different Francisella subspecies where in the most pathogenic type A variants all genes are intact while several Tfp genes are pseudogenes in the less pathogenic type B strains. This suggests that there has been a selection for expression of Tfp with different properties in the different subspecies. There is also a possibility that the genetic differences reflect adaptation to different environmental niches of the subspecies and plays a role in transmission of tularemia. This is also in line with recent findings where Tfp pilins are found to be glycosylated which could reflect a role for Tfp in the environment to promote survival and transmission. We are still far from understanding the role of Tfp in virulence and transmission of tularemia, but with the genomic information and genetic tools available we are in a good position to address these issues in the future

The type IV pilin, PilA, is required for full virulence of Francisella tularensis subspecies tularensis

Published onlineJournal ArticleThis is the final version of the article. Available from BioMed Central via the DOI in this record.BACKGROUND: All four Francisella tularensis subspecies possess gene clusters with potential to express type IV pili (Tfp). These clusters include putative pilin genes, as well as pilB, pilC and pilQ, required for secretion and assembly of Tfp. A hallmark of Tfp is the ability to retract the pilus upon surface contact, a property mediated by the ATPase PilT. Interestingly, out of the two major human pathogenic subspecies only the highly virulent type A strains have a functional pilT gene. RESULTS: In a previous study, we were able to show that one pilin gene, pilA, was essential for virulence of a type B strain in a mouse infection model. In this work we have examined the role of several Tfp genes in the virulence of the pathogenic type A strain SCHU S4. pilA, pilC, pilQ, and pilT were mutated by in-frame deletion mutagenesis. Interestingly, when mice were infected with a mixture of each mutant strain and the wild-type strain, the pilA, pilC and pilQ mutants were out-competed, while the pilT mutant was equally competitive as the wild-type. CONCLUSIONS: This suggests that expression and surface localisation of PilA contribute to virulence in the highly virulent type A strain, while PilT was dispensable for virulence in the mouse infection model

Identification of new virulence factors in Francisella tularensis

Francisella tularensis, the causative agent of tularemia, is a highly virulent bacterium with an infection dose of less than ten bacteria. The ability of a pathogen to cause infection relies on different virulence mechanisms, but in Francisella tularensis relatively few virulence factors are known. Two F. tularensis subspecies are virulent in humans; the highly virulent subspecies tularensis, also referred to as type A, and the less virulent subspecies holarctica, also called type B. The aim of this thesis has been to improve the knowledge regarding the ability of Francisella to cause disease, with the emphasis on surface located and membrane associated proteins and structures. In addition I have also investigated how virulence is regulated by studying the role of the small RNA chaperone, Hfq. The genome of Francisella appears to encode few regulatory genes. In my work I found that Hfq has an important role in regulation of virulence associated genes in Francisella. Similar to what has been found in other pathogens, Hfq functions in negative regulation, and this is the first time a negative regulation has been described for genes in the Francisella pathogenicity island. Another protein with a key role in virulence is a homologue to a disulphide oxidoreductase, DsbA, which was identified as an outer membrane lipoprotein in Francisella. A dsbA mutant was found to be severely attenuated for virulence and also induced protection against wild-type infections, thus making it a candidate for exploration as a new live vaccine. Additional genes with homology to known virulence determinants include a type IV pilin system. The pilin homologue, PilA, was identified to be required for full virulence in both type A and type B strains. In addition, genes involved in pili assembly and secretion, pilC and pilQ, were also found to be virulence associated in the type A strain. In summary, dsbA, hfq and type IV pili associated genes were indentified to be virulence determinants in F. tularensis. DsbA is a potential target for drug development and a dsbA mutant a candidate for a new live vaccine strain. Furthermore the identification of Hfq as a novel regulatory factor opens new insights into the virulence regulatory network in Francisella

Evaluating hydrogen gas transport in pipelines: Current state of numerical and experimental methodologies

This review article provides a comprehensive overview of the fundamentals, modelling approaches, experimental studies, and challenges associated with hydrogen gas flow in pipelines. It elucidates key aspects of hydrogen gas flow, including density, compressibility factor, and other relevant properties crucial for understanding its behavior in pipelines. Equations of state are discussed in detail, highlighting their importance in accurately modeling hydrogen gas flow. In the subsequent sections, one-dimensional and three-dimensional modelling techniques for gas distribution networks and localized flow involving critical components are explored. Emphasis is placed on transient flow, friction losses, and leakage characteristics, shedding light on the complexities of hydrogen pipeline transportation. Experimental studies investigating hydrogen pipeline transportation dynamics are outlined, focusing on the impact of leakage on surrounding environments and safety parameters. The challenges and solutions associated with repurposing natural gas pipelines for hydrogen transport are discussed, along with the influence of pipeline material on hydrogen transportation. Identified research gaps underscore the need for further investigation into areas such as transient flow behavior, leakage mitigation strategies, and the development of advanced modelling techniques. Future perspectives address the growing demand for hydrogen as a clean energy carrier and the evolving landscape of hydrogen-based energy systems.Full text license: CC BY 4.0;</p

Hfq, a Novel Pleiotropic Regulator of Virulence-Associated Genes in Francisella tularensis▿

Francisella tularensis is a highly infectious pathogen that infects animals and humans, causing tularemia. The ability to replicate within macrophages is central for virulence and relies on expression of genes located in the Francisella pathogenicity island (FPI), as well as expression of other genes. Regulation of FPI-encoded virulence gene expression in F. tularensis involves at least four regulatory proteins and is not fully understood. Here we studied the RNA-binding protein Hfq in F. tularensis and particularly the role that it plays as a global regulator of gene expression in stress tolerance and pathogenesis. We demonstrate that Hfq promotes resistance to several cellular stresses (including osmotic and membrane stresses). Furthermore, we show that Hfq is important for the ability of the F. tularensis vaccine strain LVS to induce disease and persist in organs of infected mice. We also demonstrate that Hfq is important for stress tolerance and full virulence in a virulent clinical isolate of F. tularensis, FSC200. Finally, microarray analyses revealed that Hfq regulates expression of numerous genes, including genes located in the FPI. Strikingly, Hfq negatively regulates only one of two divergently expressed putative operons in the FPI, in contrast to the other known regulators, which regulate the entire FPI. Hfq thus appears to be a new pleiotropic regulator of virulence in F. tularensis, acting mostly as a repressor, in contrast to the other regulators identified so far. Moreover, the results obtained suggest a novel regulatory mechanism for a subset of FPI genes

Direct repeat-mediated deletion of a type IV pilin gene results in major virulence attenuation of Francisella tularensis.

Francisella tularensis, the causative agent of tularaemia, is a highly infectious and virulent intracellular pathogen. There are two main human pathogenic subspecies, Francisella tularensis ssp. tularensis (type A), and Francisella tularensis ssp. holarctica (type B). So far, knowledge regarding key virulence determinants is limited but it is clear that intracellular survival and multiplication is one major virulence strategy of Francisella. In addition, genome sequencing has revealed the presence of genes encoding type IV pili (Tfp). One genomic region encoding three proteins with signatures typical for type IV pilins contained two 120 bp direct repeats. Here we establish that repeat-mediated loss of one of the putative pilin genes in a type B strain results in severe virulence attenuation in mice infected by subcutaneous route. Complementation of the mutant by introduction of the pilin gene in cis resulted in complete restoration of virulence. The level of attenuation was similar to that of the live vaccine strain and this strain was also found to lack the pilin gene as result of a similar deletion event mediated by the direct repeats. Presence of the pilin had no major effect on the ability to interact, survive and multiply inside macrophage-like cell lines. Importantly, the pilin-negative strain was impaired in its ability to spread from the initial site of infection to the spleen. Our findings indicate that this putative pilin is critical for Francisella infections that occur via peripheral routes

Oral health and obesity indicators

<p>Abstract</p> <p>Background</p> <p>In western Sweden, the aim was to study the associations between oral health variables and total and central adiposity, respectively, and to investigate the influence of socio-economic factors (SES), lifestyle, dental anxiety and co-morbidity.</p> <p>Methods</p> <p>The subjects constituted a randomised sample from the 1992 data collection in the Prospective Population Study of Women in Gothenburg, Sweden (n = 999, 38- > =78 yrs). The study comprised a clinical and radiographic examination, together with a self-administered questionnaire. Obesity was defined as body mass index (BMI) > =30 kg/m², waist-hip ratio (WHR) > =0.80, and waist circumference >0.88 m. Associations were estimated using logistic regression including adjustments for possible confounders.</p> <p>Results</p> <p>The mean BMI value was 25.96 kg/m², the mean WHR 0.83, and the mean waist circumference 0.83 m. The number of teeth, the number of restored teeth, xerostomia, dental visiting habits and self-perceived health were associated with both total and central adiposity, independent of age and SES. For instance, there were statistically significant associations between a small number of teeth (<20) and obesity: BMI (OR 1.95; 95% CI 1.40-2.73), WHR (1.67; 1.28-2.19) and waist circumference (1.94; 1.47-2.55), respectively. The number of carious lesions and masticatory function showed no associations with obesity. The obesity measure was of significance, particularly with regard to behaviour, such as irregular dental visits, with a greater risk associated with BMI (1.83; 1.23-2.71) and waist circumference (1.96; 1.39-2.75), but not with WHR (1.29; 0.90-1.85).</p> <p>Conclusions</p> <p>Associations were found between oral health and obesity. The choice of obesity measure in oral health studies should be carefully considered.</p