Development and Comparison of a Panel of Modified CS17 Fimbrial Tip Adhesin Proteins as Components for an Adhesin-Based Vaccine against Enterotoxigenic Escherichia coli

Enterotoxigenic Escherichia coli (ETEC) is a leading cause of diarrhea in travelers and children in resource-limited countries. ETEC colonization factors, fimbrial tip adhesins and enterotoxins are key virulence factors, and thus have been studied as vaccine candidates. Some prevalent colonization factors, including CFA/I and CS17, belong to the class 5 family. We previously found that passive oral administration of hyperimmune bovine colostral IgG (bIgG) raised against dscCfaE (donor strand complemented CFA/I tip adhesin) protected volunteers against CFA/I+ ETEC challenge, while anti-dscCsbD bIgG (CS17 tip adhesin) did not confer protection. These findings led us to develop and optimize a panel of alternative CsbD-based vaccine candidates based on allele matching and in silico protein engineering. Physicochemical characterizations revealed that an optimized vaccine candidate dscCsbDLSN139(P218A/G3) had the greatest thermal stability among the six tested dscCsbD adhesins, whereas the overall secondary structures and solubility of these adhesins had no obvious differences. Importantly, dscCsbDLSN139(P218A/G3) elicited significantly higher CS17+ ETEC hemagglutination inhibition titers in sera from mice intranasally immunized with the panel of dscCsbD adhesins, while no significant difference was observed among heterologous neutralizing titers. Our results strongly advocate for the incorporation of these modifications into a new generation of CsbD-based ETEC vaccine candidates.

Comprehensive genetic analysis of adhesin proteins and their role in virulence ofCandida albicans

Candida albicans is a microbial fungus that exists as a commensal member of the human microbiome and an opportunistic pathogen. Cell surface-associated adhesin proteins play a crucial role in C. albicans’ ability to undergo cellular morphogenesis, develop robust biofilms, colonize, and cause infection in a host. However, a comprehensive analysis of the role and relationships between these adhesins has not been explored. We previously established a CRISPR-based platform for efficient generation of single- and double-gene deletions in C. albicans, which was used to construct a library of 144 mutants, comprising 12 unique adhesin genes deleted singly, and every possible combination of double deletions. Here, we exploit this adhesin mutant library to explore the role of adhesin proteins in C. albicans virulence. We perform a comprehensive, high-throughput screen of this library, using Caenorhabditis elegans as a simplified model host system, which identified mutants critical for virulence and significant genetic interactions. We perform follow-up analysis to assess the ability of high- and low-virulence strains to undergo cellular morphogenesis and form biofilms in vitro, as well as to colonize the C. elegans host. We further perform genetic interaction analysis to identify novel significant negative genetic interactions between adhesin mutants, whereby combinatorial perturbation of these genes significantly impairs virulence, more than expected based on virulence of the single mutant constituent strains. Together, this study yields important new insight into the role of adhesins, singly and in combinations, in mediating diverse facets of virulence of this critical fungal pathogen.

Systematic characterization and genetic interaction analysis of adhesins in Candida albicans virulence

Candida albicans is a microbial fungus that exists as a commensal member of the human microbiome and an opportunistic pathogen. Cell surface-associated adhesin proteins play a crucial role in C. albicans’ ability to undergo cellular morphogenesis, develop robust biofilms, colonize, and cause infection in a host. However, a comprehensive analysis of the role and relationships between these adhesins has not been explored. We previously established a CRISPR-based platform for efficient generation of single- and double-gene deletions in C. albicans, which was used to construct a library of 144 mutants, comprising 12 unique adhesin genes deleted singly, or in every possible combination of double deletions. Here, we exploit this adhesin mutant library to explore the role of adhesin proteins in C. albicans virulence. We perform a comprehensive, high-throughput screen of this library, using Caenorhabditis elegans as a simplified model host system, which identified mutants critical for virulence and significant genetic interactions. We perform follow-up analysis to assess the ability of high- and low-virulence strains to undergo cellular morphogenesis and form biofilms in vitro, as well as to colonize the C. elegans host. We further perform genetic interaction analysis to identify novel significant negative genetic interactions between adhesin mutants, whereby combinatorial perturbation of these genes significantly impairs virulence, more than expected based on virulence of the single mutant constituent strains. Together, this yields important new insight into the role of adhesins, singly and in combinations, in mediating diverse facets of virulence of this critical fungal pathogen.SummaryCandida albicans is a human fungal pathogen and cause of life-threatening systemic infections. Cell surface-associated adhesins play a central role in this pathogen’s ability to establish infection. Here, we provide a comprehensive analysis of adhesin factors, and their role in fungal virulence. Exploiting a high-throughput workflow, we screened an adhesin mutant library using C. elegans as a simple model host, and identified mutants and genetic interactions involved in virulence. We found that adhesin mutants are impaired in in vitro pathogenicity, irrespective of their virulence. Together, this work provides new insight into the role of adhesin factors in mediating fungal virulence.

The Role of Pili Protein 38,6 kDa Klebsiella pneumoniae as a Hemagglutinin and Adhesin Protein which Serves as a Virulence Factor

Klebsiella pneumoniae is an infectious bacteria in various parts of the body. The presence of proteins in pili that will bind to cell surface receptors (adhesin proteins) and cell membrane sugar molecules (hemagglutinin proteins) is a very influential factor in the ability of K. pneumoniae to enter the human body. The purpose of this study was to determine the role of pili 38.6 kDa K. pneumoniae protein as a hemagglutinin and adhesin protein which functions as a virulence factor. This type of research is pure experimental research using the electrophoresis method (SDS-PAGE) to obtain protein. The protein obtained was then tested for hemagglutination and adhesion test using erythrocyte cells and enterocytes of BALB / C mice to determine their role as hemagglutinin and adhesin proteins. Conclusion of this study pili protein 38.6 kDa Klebsiella pneumoniae acts as a hemagglutinin and adhesion protein which functions as a virulence factor. Keywords: Klebsiella pneumonia, 38,6 kDa Protein, Hemagglutinin, Adhesin

Role of Evolutionary Selection Acting on Vaccine Antigens in the Re-Emergence of Bordetella Pertussis

Pertussis (“whooping cough”) is a re-emerging disease with increasing incidence among fully vaccinated individuals. We explored the genetic diversity of five Bordetella pertussis proteins used to generate the subunit vaccine across ancestral and newly emergent strains using immunoinformatics and evolutionary selection measurements. The five subunits of pertussis toxin (Ptx1–Ptx5) were highly conserved with regard to sequence, predicted structure, predicted antigenicity, and were under purifying selection. In contrast, the adhesin proteins pertactin (Prn) and filamentous hemagglutinin (FHA) were under statistically significant (p < 0.01) diversifying selection. Most heavily diversified sites of each protein fell within antigenic epitopes, and the functional adhesin motifs were conserved. Protein secondary structure was conserved despite sequence diversity for FHA but was changeable in Prn. These findings suggest that subunit vaccine-derived immunity does not impact Ptx1–Ptx5 but may apply evolutionary pressure to Prn and FHA to undergo diversifying selection. These findings offer further insight into the emergence of vaccine-resistant strains of B. pertussis.

BIOFILMS OF PATHOGENIC BURKHOLDERIA AND THEIR ROLE IN RESISTANCE TO ANTIBIOTICS

The review contains the current knowledge on the main issues of Burkholderia pseudomallei and Burkholderia mallei biofilm formation. The role ofknown structural elements of Burkholderia cells (flagella, type IV pili, LPS), as well as autotransporter adhesin proteins in the attachment of bacteria to surfaces, the formation of microcolonies and biofilm is described. The review also includes information of genetic regulatory mechanisms (QS-systems, RpoE-sigma factor, c-di-GMP, two-component signal transduction system), differentially expressed genes related to the formation of B. pseudomallei biofilm, role ofbiofilms in the virulence and resistance to antibiotics of pathogenic Burkholderia and their significance for the chronic processes and recurrent course of melioidosis and glanders.

Mechanism of a cytosolic O-glycosyltransferase essential for the synthesis of a bacterial adhesion protein

O-glycosylation of Ser and Thr residues is an important process in all organisms, which is only poorly understood. Such modification is required for the export and function of adhesin proteins that mediate the attachment of pathogenic Gram-positive bacteria to host cells. Here, we have analyzed the mechanism by which the cytosolic O-glycosyltransferase GtfA/B of Streptococcus gordonii modifies the Ser/Thr-rich repeats of adhesin. The enzyme is a tetramer containing two molecules each of GtfA and GtfB. The two subunits have the same fold, but only GtfA contains an active site, whereas GtfB provides the primary binding site for adhesin. During a first phase of glycosylation, the conformation of GtfB is restrained by GtfA to bind substrate with unmodified Ser/Thr residues. In a slow second phase, GtfB recognizes residues that are already modified with N-acetylglucosamine, likely by converting into a relaxed conformation in which one interface with GtfA is broken. These results explain how the glycosyltransferase modifies a progressively changing substrate molecule.

A.actinomycetemcomitans adhesi protein increasing IL-8 titre in heart of wistar rat with aggressive periodontitis (Protein adhesin dari A.actinomycetemcomitans meningkatkantiter IL -8 di dalamjantungtikuswistar denganperiodontitis agresif)

Cardiovascular and periodontal diseases are common inflammatory conditions in the human population. Tlymphocytesparticipateinthepathogenesis and inflammatory events of atherosclerosis. These immune cells enter theinflamed artery wall and join macrophages via a number of interferon-c-inducible chemokines. Chemokines (IL-8),cytokines, and growth factors also participate in this process. The interaction between interleukin- 8 and its receptor,CXCR2, can also contribute to lesion formation in mice. The main causes of aggressive periodontitis isActinobacillusactinomycetemcomitans. Previous studies have proven that adhesin protein with 24 kDa molecular weight from A.actinomycetemcomitans is a specific adhesin, this adhesin proteins play a role in the adhesion process on host. thiskind adhesion in the epithelial attachment would lead to colonization and invasion ofA.actinomycetemcomitans thatwill stimulate the host immune response. This study aimed to analyze the influence of induction 24 kDaA.actinomycetemcomitans adhesin protein to the titre of IL-8 in heart of Wistar rat with aggressive periodontitis usingElisa method to measure and analyze the titre of IL-8. After analyzed with analysis of variance, showed significantdifferences of IL-8 titre in the control group and the group with the induction byA.actinomycetemcomitans,A.actinomycetemcomitans plus 24 kDa A.actinomycetemcomitans adhesin protein, and only with 24 kDaA.actinomycetemcomitans adhesin protein. It can be concluded that A.actinomycetemcomitans adhesin protein with 24kDa molecular weight has a role in increasing of IL-8 titre in heart wistar rat with aggressive periodontitis.

Cell Signals, Cell Contacts, and the Organization of Yeast Communities

ABSTRACT Even relatively simple species have evolved mechanisms to organize individual organisms into communities, such that the fitness of the group is greater than the fitness of isolated individuals. Within the fungal kingdom, the ability of many yeast species to organize into communities is crucial for their growth and survival, and this property has important impacts both on the economy and on human health. Over the last few years, studies of Saccharomyces cerevisiae have revealed several fundamental properties of yeast communities. First, strain-to-strain variation in the structures of these groups is attributable in part to variability in the expression and functions of adhesin proteins. Second, the extracellular matrix surrounding these communities can protect them from environmental stress and may also be important in cell signaling. Finally, diffusible signals between cells contribute to community organization so that different regions of a community express different genes and adopt different cell fates. These findings provide an arena in which to view fundamental mechanisms by which contacts and signals between individual organisms allow them to assemble into functional communities.