Functional Amyloids in Pseudomonas aeruginosa Are Essential for the Proteome Modulation That Leads to Pathoadaptation in Pulmonary Niches.

Persistence and survival of Pseudomonas aeruginosa in chronic lung infections is closely linked to the biofilm lifestyle. One biofilm component, functional amyloid of P. aeruginosa (Fap), imparts structural adaptations for biofilms; however, the role of Fap in pathogenesis is still unclear. Conservation of the fap operon encoding Fap and P. aeruginosa being an opportunistic pathogen of lung infections prompted us to explore its role in lung infection. We found that Fap is essential for establishment of lung infection in rats, as its genetic exclusion led to mild focal infection with quick resolution. Moreover, without an underlying cystic fibrosis (CF) genetic disorder, overexpression of Fap reproduced the CF pathotype. The molecular basis of Fap-mediated pulmonary adaptation was explored through surface-associated proteomics in vitro. Differential proteomics positively associated Fap expression with activation of known proteins related to pulmonary pathoadaptation, attachment, and biofilm fitness. The aggregative bacterial phenotype in the pulmonary niche correlated with Fap-influenced activation of biofilm sustainability regulators and stress response regulators that favored persistence-mediated establishment of pulmonary infection. Fap overexpression upregulated proteins that are abundant in the proteome of P. aeruginosa in colonizing CF lungs. Planktonic lifestyle, defects in anaerobic pathway, and neutrophilic evasion were key factors in the absence of Fap that impaired establishment of infection. We concluded that Fap is essential for cellular equilibration to establish pulmonary infection. Amyloid-induced bacterial aggregation subverted the immune response, leading to chronic infection by collaterally damaging tissue and reinforcing bacterial persistence. IMPORTANCE Pseudomonas aeruginosa is inextricably linked with chronic lung infections. In this study, the well-conserved Fap operon was found to be essential for pathoadaptation in pulmonary infection in a rat lung model. Moreover, the presence of Fap increased pathogenesis and biofilm sustainability by modulating bacterial physiology. Hence, a pathoadaptive role of Fap in pulmonary infections can be exploited for clinical application by targeting amyloids. Furthermore, genetic conservation and extracellular exposure of Fap make it a commendable target for such interventions.

Designing multi-epitope vaccine candidates against functional amyloids in Pseudomonas aeruginosa through immunoinformatic and structural bioinformatics approach.

Pseudomonas aeruginosa (P. aeruginosa) displays high drug resistance and biofilm-mediated adaptability, which makes its infections difficult to treat. Alternative intervention methods and targets have made such infections treatment manageable. One of the biofilm components, functional amyloids of Pseudomonas (Fap) is correlated positively with virulence and mucoidy phenotype found in infection in cystic fibrosis (CF) patients. Extracellular accessibility, conservation across P. aeruginosa isolates and linkage with lung infections phenotype in CF patients, makes Fap a promising intervention target. Furthermore, the reported effect of bacterial amyloid on neuronal function and immune response makes it a targetable candidate. In the current study, Fap C protein and its immediate interactions were explored to extract antigenic T-cell and B-cell epitopes. A combination of epitopes and peptide adjuvants has been linked to derive vaccine candidate structures. The vaccine candidates were validated for antigenicity, allergenicity, physiochemical properties, stability and interactions with TLRs and MHC alleles. Immunosimulation studies have demonstrated that vaccines elicit Th1 dominated response, which can assist in good prognosis of infection in CF patients.

Motifs and interface amino acid-mediated regulation of amyloid biogenesis in microbes to humans: potential targets for intervention.

Amyloids are linked to many debilitating diseases in mammals. Some organisms produce amyloids that have a functional role in the maintenance of their biological processes. Microbes utilize functional bacterial amyloids (FuBA) for pathogenicity and infections. Amyloid biogenesis is regulated differentially in various systems to avoid its toxic accumulation. A familiar feature in the process of amyloid biogenesis from humans to microbes is its regulation by protein-protein interactions (PPI). The spatial arrangement of amino acid residues in proteins generates topologies like flat interface and linear motif, which participate in protein interactions. Motifs and interface residue-mediated interactions have a direct or an indirect impact on amyloid secretion and assembly. Some motifs undergo post-translational modifications (PTM), which effects interactions and dynamics of the amyloid biogenesis cascade. Interaction-induced local changes stimulate global conformational transitions in the PPI complex, which indirectly affects amyloid formation. Perturbation of such motifs and interface residues results in amyloid abolishment. Interface residues, motifs and their respective interactive protein partners could serve as potential targets for intervention to inhibit amyloid biogenesis.

Genome analyses of blaNDM-4 carrying ST 315 Escherichia coli isolate from sewage water of one of the Indian hospitals.

BACKGROUND: Emergence of carbapenem resistant Escherichia coli pathovars and their environmental dissemination are alarming problems. E. coli isolated from sewage water of hospital setting conferred a high resistance towards ß-lactams, particularly towards carbapenem. This prompted us to perform whole genome sequence analysis to investigate the antimicrobial determinants, pathogenicity status and mobile genetic elements associated with resistance genes. RESULTS: To the best of our knowledge this is the first report of ST 315 carrying NDM-4 from India. The genome analysis has revealed the unknown characteristics associated with this sequence type (ST 315) like resistance and virulence factors. Based on virulence markers, its pathotype was identified as ExPEC. Furthermore, a mobile plasmid with multiple ß-lactamases genes and clinically relevant resistance markers was detected. Phylogenetic analysis of Inc F plasmids sequences carrying ESBLs and NDM variants, revealed un-relatedness in these plasmids due to their varying size and backbone sequences. CONCLUSIONS: Presence of carbapenem resistant E. coli ST 315 with high level antibiotic resistance, near hospital environment is an alarming situation in context to its spread. WGS based analyses have provided details on virulence and resistance status which could overcome the lack of information available on ST 315, globally. This could further help in its quick detection and control in clinical settings.

Draft Genome Sequence of the First NDM-4-Producing Escherichia coli Strain (AK1), Isolated from Sewage Water of a North Indian Hospital.

We report here the draft genome sequence of the first isolated NDM-4-producing Escherichia coli strain, isolated from sewage water at a North Indian hospital. The genome has an assembly size of 5,076,053 bp, arranged in 129 contigs, with 5,271 genes and a G+C content of 50.47%.