Predictability studies of coastal marine ecosystem behavior

The study presented in this thesis is principally meant to analyze the genericity of a deterministic, comprehensive marine ecosystem model in combination with various refined representations of hydrodynamical processes, and to evaluate the potential predictability skills of this combined modelling system with specific applications in two rather different coastal basins. This objective has been realized by first developing a modular coupling interface between the Princeton Ocean Model (POM) and the European Regional Seas Ecosystem Model (ERSEM), called High-Resolution OpenSESAME POM ERSEM (HiROPE). Secondly, this model framework, embedding a composite of 'complex' conceptual principles of the functioning of the main biogeochemical processes, has been applied to substantially different marine systems, the Baltic proper and the northern Adriatic Sea. The generic biological first principles of the ERSEM ecosystem model have been throughly controlled for consistency, and a suitable mathematical syntax has been defined in order to accomodate the various biogeochemical cycles of the resolved elements. The model has been specifically applied in the chosen basins with different temporal and spatial resolutions: a one-dimensional (vertical, 1D-V), climatological implementation in the northern Adriatic Sea; a 1D-V implementation in the Baltic proper with realistic forcing functions in the period 1979-1991 and a fully three-dimensional, high-frequency realistic implementation in the northern Adriatic Sea (October 1995). General conclusions are that the representation of hydrodynamical variability, the definition and resolution of boundary processes, the introduction of new source terms or the implementation of new biological state variables, affect the predictability of the system behavior more than the utilization of incomplete initial conditions of biological variables in a complex comprehensive ecosystem model

Insights into a Multidrug Resistant Escherichia coli Pathogen of the Globally Disseminated ST131 Lineage: Genome Analysis and Virulence Mechanisms

Escherichia coli strains causing urinary tract infection (UTI) are increasingly recognized as belonging to specific clones. E. coli clone O25b:H4-ST131 has recently emerged globally as a leading multi-drug resistant pathogen causing urinary tract and bloodstream infections in hospitals and the community. While most molecular studies to date examine the mechanisms conferring multi-drug resistance in E. coli ST131, relatively little is known about their virulence potential. Here we examined E. coli ST131 clinical isolates from two geographically diverse collections, one representing the major pathogenic lineages causing UTI across the United Kingdom and a second representing UTI isolates from patients presenting at two large hospitals in Australia. We determined a draft genome sequence for one representative isolate, E. coli EC958, which produced CTX-M-15 extended-spectrum β-lactamase, CMY-23 type AmpC cephalosporinase and was resistant to ciprofloxacin. Comparative genome analysis indicated that EC958 encodes virulence genes commonly associated with uropathogenic E. coli (UPEC). The genome sequence of EC958 revealed a transposon insertion in the fimB gene encoding the activator of type 1 fimbriae, an important UPEC bladder colonization factor. We identified the same fimB transposon insertion in 59% of the ST131 UK isolates, as well as 71% of ST131 isolates from Australia, suggesting this mutation is common among E. coli ST131 strains. Insertional inactivation of fimB resulted in a phenotype resembling a slower off-to-on switching for type 1 fimbriae. Type 1 fimbriae expression could still be induced in fimB-null isolates; this correlated strongly with adherence to and invasion of human bladder cells and bladder colonisation in a mouse UTI model. We conclude that E. coli ST131 is a geographically widespread, antibiotic resistant clone that has the capacity to produce numerous virulence factors associated with UTI

Intestinal Colonization Traits of Pandemic Multidrug-Resistant Escherichia coli ST131

Background. Epidemiological studies point to the gut as a key reservoir of multidrug resistant Escherichia coli multilocus sequence type 131 (ST131), a globally dominant pathogenic clone causing urinary tract and bloodstream infections. Here we report a detailed investigation of its intestinal lifestyle. Methods. Clinical ST131 isolates and type 1 fimbriae null mutants were assessed for colonization of human intestinal epithelia and in mouse intestinal colonization models. Mouse gut tissue underwent histologic analysis for pathology and ST131 localization. Key findings were corroborated in mucus-producing human cell lines and intestinal biopsy specimens. Results. ST131 strains adhered to and invaded human intestinal epithelial cells more than probiotic and commensal strains. The reference ST131 strain EC958 established persistent intestinal colonization in mice, and expression of type 1 fimbriae mediated higher colonization levels. Bacterial loads were highest in the distal parts of the mouse intestine and did not cause any obvious pathology. Further analysis revealed that EC958 could bind to both mucus and underlying human intestinal epithelia. Conclusions. ST131 strains can efficiently colonize the mammalian gut and persist long term. Type 1 fimbriae enhance ST131 intestinal colonization, suggesting that mannosides, currently developed as therapeutics for bladder infections and Crohn’s disease, could also be used to limit intestinal ST131 reservoirs

Incidence of multiple Herpesvirus infection in HIV seropositive patients, a big concern for Eastern Indian scenario

<p>Abstract</p> <p>Background</p> <p>Human immunodeficiency virus (HIV) infection is associated with an increased risk for human <it>herpes viruses </it>(HHVs) and their related diseases and they frequently cause disease deterioration and therapeutic failures. Methods for limiting the transmission of HHVs require a better understanding of the incidence and infectivity of oral HHVs in HIV-infected patients. This study was designed to determine the seroprevalence of human herpes viruses (CMV, HSV 2, EBV-1, VZV) antibodies and to evaluate their association with age, sex as well as other demographic and behavioral factors.</p> <p>Results</p> <p>A study of 200 HIV positive patients from Eastern India attending the Calcutta Medical College Hospital, Kolkata, West Bengal, Apex Clinic, Calcutta Medical College Hospital and ART Center, School of Tropical Medicine, Kolkata, West Bengal was done. Serum samples were screened for antibodies to the respective viruses using the indirect ELISA in triplicates.</p> <p><it>CytoMegalo virus </it>(CMV), <it>Herpes Simplex virus </it>type 2 (HSV-2), <it>Varicella Zoster virus </it>(VZV), and <it>Epstein Barr virus </it>(EBV-1) were detected in 49%, 47%, 32.5%, and 26% respectively.</p> <p>Conclusion</p> <p>This study has contributed baseline data and provided insights in viral OI and HIV co-infection in Eastern India. This would undoubtedly serve as a basis for further studies on this topic.</p

Role of capsule and O antigen in the virulence of uropathogenic Escherichia coli

Urinary tract infection (UTI) is one of the most common bacterial infections in humans, with uropathogenic Escherichia coli (UPEC) the leading causative organism. UPEC has a number of virulence factors that enable it to overcome host defenses within the urinary tract and establish infection. The O antigen and the capsular polysaccharide are two such factors that provide a survival advantage to UPEC. Here we describe the application of the rpsL counter selection system to construct capsule (kpsD) and O antigen (waaL) mutants and complemented derivatives of three reference UPEC strains: CFT073 (O6:K2:H1), RS218 (O18:K1:H7) and 1177 (O1:K1:H7). We observed that while the O1, O6 and O18 antigens were required for survival in human serum, the role of the capsule was less clear and linked to O antigen type. In contrast, both the K1 and K2 capsular antigens provided a survival advantage to UPEC in whole blood. In the mouse urinary tract, mutation of the O6 antigen significantly attenuated CFT073 bladder colonization. Overall, this study contrasts the role of capsule and O antigen in three common UPEC serotypes using defined mutant and complemented strains. The combined mutagenesis-complementation strategy can be applied to study other virulence factors with complex functions both in vitro and in vivo

Conserved FimH mutations in the global Escherichia coli ST131 multi-drug resistant lineage weaken interdomain interactions and alter adhesin function

The binding of the type 1 fimbrial adhesin FimH to mannosylated receptors is allosterically regulated to enhance the fitness of uropathogenic Escherichia coli (UPEC) during urinary tract infection (UTI). Mutations in the two FimH domains (pilin and lectin) located outside the mannose binding pocket have been shown to influence mannose binding affinity, yet the details of the allostery mechanism are not fully elucidated. Here we characterised different FimH conformational states (termed low-affinity tense and high-affinity relaxed conformations) of natural FimH variants using molecular dynamics (MD) simulation techniques and report key structural dynamics differences between them. The clinically dominant FimH30 variant from the pandemic multidrug resistant E. coli ST131 lineage contains an R166H mutation that weakens FimH interdomain interactions and allows enhanced mannose interactions with pre-existing high-affinity relaxed conformations. When expressed in an isogenic ST131 strain background, FimH30 mediated high human cell adhesion and invasion, and enhanced biofilm formation over other variants. Collectively, our computational and experimental findings support a model of FimH protein allostery that is mediated by shifts in the pre-existing conformational equilibrium of FimH, additional to the sequential step-wise process of structural perturbations transmitted from one site to another within the protein. Importantly, it is the first study to shed light into how natural mutations in a clinically dominant FimH variant influence the protein's conformational landscape optimising its function for ST131 fitness at intestinal and extraintestinal niches.</p

F9 Fimbriae of uropathogenic Escherichia coli are expressed at low temperature and recognise Galbeta1-3GlcNAc-containing glycans

Uropathogenic Escherichia coli (UPEC) is the leading causative agent of urinary tract infections (UTI) in the developed world. Among the major virulence factors of UPEC, surface expressed adhesins mediate attachment and tissue tropism. UPEC strains typically possess a range of adhesins, with type 1 fimbriae and P fimbriae of the chaperone-usher class the best characterised. We previously identified and characterised F9 as a new chaperone-usher fimbrial type that mediates biofilm formation. However, the regulation and specific role of F9 fimbriae remained to be determined in the context of wild-type clinical UPEC strains. In this study we have assessed the distribution and genetic context of the f9 operon among diverse E. coli lineages and pathotypes and demonstrated that f9 genes are significantly more conserved in a UPEC strain collection in comparison to the well-defined E. coli reference (ECOR) collection. In the prototypic UPEC strain CFT073, the global regulator protein H-NS was identified as a transcriptional repressor of f9 gene expression at 37°C through its ability to bind directly to the f9 promoter region. F9 fimbriae expression was demonstrated at 20°C, representing the first evidence of functional F9 fimbriae expression by wild-type E. coli. Finally, glycan array analysis demonstrated that F9 fimbriae recognise and bind to terminal Galβ1-3GlcNAc structures

Fitness of CFT073 capsule (<i>kpsD</i>) and O antigen (<i>waaL</i>) mutants during <i>in vivo</i> competition with CFT073 WT and corresponding complemented (<i>kpsD</i>^C, <i>waaL</i>^C) strains.

<p>C57B/L6 mice were infected transurethally with a 1∶1 mixture of CFT073<i>kpsD</i> and CFT073 WT or CFT073<i>kpsD</i>^C; CFT073<i>waaL</i> and CFT073 WT or CFT073<i>waaL</i>^C. (A) Each marker represents LOG₁₀ total CFU recovered from each mouse per 0.1 g of bladder tissue. Lines connect data points for the same mouse, and horizontal bars represent median values. (B) Each marker represents the LOG₁₀ competitive index calculated for each individual mouse; competitive indices are the ratio of the mutant 0.1 g of bladder tissue to that of WT or complemented strain. Dashed lines represent hypothetical competitive index of 1 (LOG₁₀1 = 0), which indicates no difference in fitness between the two strains. Horizontal bars represent group medians, and each competition group had 6 to 8 mice. All mutants were significantly outcompeted by CFT073 WT or their respective complemented strains for bladder colonization (<i>P</i><0.05). The O6 antigen was significantly more important than the K2 capsular antigen for bladder colonization (<i>P<</i>0.05).</p