Cyanobacterial Secretion Systems: Understanding Fundamental Mechanisms Toward Technological Applications

Avian pathogenic Escherichia coli (APEC) causes colibacillosis in avian species, and recent reports have suggested APEC as a potential foodborne zoonotic pathogen. Herein, we discuss the virulence and pathogenesis factors of APEC, review the zoonotic potential, provide the current status of antibiotic resistance and progress in vaccine development, and summarize the alternative control measures being investigated. In addition to the known virulence factors, several other factors including quorum sensing system, secretion systems, two-component systems, transcriptional regulators, and genes associated with metabolism also contribute to APEC pathogenesis. The clear understanding of these factors will help in developing new effective treatments. The APEC isolates (particularly belonging to ST95 and ST131 or O1, O2, and O18) have genetic similarities and commonalities in virulence genes with human uropathogenic E. coli (UPEC) and neonatal meningitis E. coli (NMEC) and abilities to cause urinary tract infections and meningitis in humans. Therefore, the zoonotic potential of APEC cannot be undervalued. APEC resistance to almost all classes of antibiotics, including carbapenems, has been already reported. There is a need for an effective APEC vaccine that can provide protection against diverse APEC serotypes. Alternative therapies, especially the virulence inhibitors, can provide a novel solution with less likelihood of developing resistance.

Download Full-text

Clonal Clusters, Molecular Resistance Mechanisms and Virulence Factors of Gram-Negative Bacteria Isolated from Chronic Wounds in Ghana

Antibiotics ◽

10.3390/antibiotics10030339 ◽

2021 ◽

Vol 10 (3) ◽

pp. 339

Author(s):

Denise Dekker ◽

Frederik Pankok ◽

Thorsten Thye ◽

Stefan Taudien ◽

Kwabena Oppong ◽

...

Keyword(s):

Molecular Epidemiology ◽

Virulence Factors ◽

Iron Uptake ◽

Chronic Wounds ◽

Sub Saharan Africa ◽

Gram Negative Bacteria ◽

Beta Lactamase ◽

Secretion Systems ◽

Gram Negative ◽

E Coli

Wound infections are common medical problems in sub-Saharan Africa but data on the molecular epidemiology are rare. Within this study we assessed the clonal lineages, resistance genes and virulence factors of Gram-negative bacteria isolated from Ghanaian patients with chronic wounds. From a previous study, 49 Pseudomonas aeruginosa, 21 Klebsiellapneumoniae complex members and 12 Escherichia coli were subjected to whole genome sequencing. Sequence analysis indicated high clonal diversity with only nine P. aeruginosa clusters comprising two strains each and one E. coli cluster comprising three strains with high phylogenetic relationship suggesting nosocomial transmission. Acquired beta-lactamase genes were observed in some isolates next to a broad spectrum of additional genetic resistance determinants. Phenotypical expression of extended-spectrum beta-lactamase activity in the Enterobacterales was associated with blaCTX-M-15 genes, which are frequent in Ghana. Frequently recorded virulence genes comprised genes related to invasion and iron-uptake in E. coli, genes related to adherence, iron-uptake, secretion systems and antiphagocytosis in P. aeruginosa and genes related to adherence, biofilm formation, immune evasion, iron-uptake and secretion systems in K. pneumonia complex. In summary, the study provides a piece in the puzzle of the molecular epidemiology of Gram-negative bacteria in chronic wounds in rural Ghana.

Download Full-text

Novel Template Plasmids pCyaA’-Kan and pCyaA’-Cam for Generation of Unmarked Chromosomal cyaA’ Translational Fusion to T3SS Effectors in Salmonella

Microorganisms ◽

10.3390/microorganisms9030475 ◽

2021 ◽

Vol 9 (3) ◽

pp. 475

Author(s):

Paulina A. Fernández ◽

Marcela Zabner ◽

Jaime Ortega ◽

Constanza Morgado ◽

Fernando Amaya ◽

...

Keyword(s):

Fusion Protein ◽

Western Blot ◽

Eukaryotic Cells ◽

Intracellular Camp ◽

Gene Fusions ◽

Secretion Systems ◽

Translational Fusion ◽

Flp Recombinase ◽

Recombination System ◽

Camp Levels

The type III secretion systems (T3SS) encoded in pathogenicity islands SPI-1 and SPI-2 are key virulence factors of Salmonella. These systems translocate proteins known as effectors into eukaryotic cells during infection. To characterize the functionality of T3SS effectors, gene fusions to the CyaA’ reporter of Bordetella pertussis are often used. CyaA’ is a calmodulin-dependent adenylate cyclase that is only active within eukaryotic cells. Thus, the translocation of an effector fused to CyaA’ can be evaluated by measuring cAMP levels in infected cells. Here, we report the construction of plasmids pCyaA’-Kan and pCyaA’-Cam, which contain the ORF encoding CyaA’ adjacent to a cassette that confers resistance to kanamycin or chloramphenicol, respectively, flanked by Flp recombinase target (FRT) sites. A PCR product from pCyaA’-Kan or pCyaA’-Cam containing these genetic elements can be introduced into the bacterial chromosome to generate gene fusions by homologous recombination using the Red recombination system from bacteriophage λ. Subsequently, the resistance cassette can be removed by recombination between the FRT sites using the Flp recombinase. As a proof of concept, the plasmids pCyaA’-Kan and pCyaA’-Cam were used to generate unmarked chromosomal fusions of 10 T3SS effectors to CyaA’ in S. Typhimurium. Each fusion protein was detected by Western blot using an anti-CyaA’ monoclonal antibody when the corresponding mutant strain was grown under conditions that induce the expression of the native gene. In addition, T3SS-1-dependent secretion of fusion protein SipA-CyaA’ during in vitro growth was verified by Western blot analysis of culture supernatants. Finally, efficient translocation of SipA-CyaA’ into HeLa cells was evidenced by increased intracellular cAMP levels at different times of infection. Therefore, the plasmids pCyaA’-Kan and pCyaA’-Cam can be used to generate unmarked chromosomal cyaA’ translational fusion to study regulated expression, secretion and translocation of Salmonella T3SS effectors into eukaryotic cells.

Download Full-text

Folding Control in the Path of Type 5 Secretion

Toxins ◽

10.3390/toxins13050341 ◽

2021 ◽

Vol 13 (5) ◽

pp. 341

Author(s):

Nathalie Dautin

Keyword(s):

Protein Folding ◽

Virulence Factors ◽

Secretion System ◽

Gram Negative Bacteria ◽

Secretion Systems ◽

Gram Negative ◽

Final Destination ◽

Cell Compartments ◽

Recent Addition ◽

Functionally Diverse

The type 5 secretion system (T5SS) is one of the more widespread secretion systems in Gram-negative bacteria. Proteins secreted by the T5SS are functionally diverse (toxins, adhesins, enzymes) and include numerous virulence factors. Mechanistically, the T5SS has long been considered the simplest of secretion systems, due to the paucity of proteins required for its functioning. Still, despite more than two decades of study, the exact process by which T5SS substrates attain their final destination and correct conformation is not totally deciphered. Moreover, the recent addition of new sub-families to the T5SS raises additional questions about this secretion mechanism. Central to the understanding of type 5 secretion is the question of protein folding, which needs to be carefully controlled in each of the bacterial cell compartments these proteins cross. Here, the biogenesis of proteins secreted by the Type 5 secretion system is discussed, with a focus on the various factors preventing or promoting protein folding during biogenesis.

Download Full-text

Type VII secretion systems: structure, functions and transport models

Nature Reviews Microbiology ◽

10.1038/s41579-021-00560-5 ◽

2021 ◽

Author(s):

Angel Rivera-Calzada ◽

Nikolaos Famelis ◽

Oscar Llorca ◽

Sebastian Geibel

Keyword(s):

Structure Functions ◽

Secretion Systems ◽

Transport Models ◽

Type Vii Secretion

Download Full-text

Systematic Detection of Large-Scale Multi-Gene Horizontal Transfer in Prokaryotes

Molecular Biology and Evolution ◽

10.1093/molbev/msab043 ◽

2021 ◽

Author(s):

Lina Kloub ◽

Sean Gosselin ◽

Matthew Fullmer ◽

Joerg Graf ◽

J Peter Gogarten ◽

...

Keyword(s):

Gene Transfer ◽

Large Scale ◽

Single Gene ◽

Gene Families ◽

Microbial Evolution ◽

Phylogenetic Distance ◽

Secretion Systems ◽

Type Iii Secretion Systems ◽

A Genome ◽

Conserved Gene

Abstract Horizontal gene transfer (HGT) is central to prokaryotic evolution. However, little is known about the “scale” of individual HGT events. In this work, we introduce the first computational framework to help answer the following fundamental question: How often does more than one gene get horizontally transferred in a single HGT event? Our method, called HoMer, uses phylogenetic reconciliation to infer single-gene HGT events across a given set of species/strains, employs several techniques to account for inference error and uncertainty, combines that information with gene order information from extant genomes, and uses statistical analysis to identify candidate horizontal multi-gene transfers (HMGTs) in both extant and ancestral species/strains. HoMer is highly scalable and can be easily used to infer HMGTs across hundreds of genomes. We apply HoMer to a genome-scale dataset of over 22000 gene families from 103 Aeromonas genomes and identify a large number of plausible HMGTs of various scales at both small and large phylogenetic distances. Analysis of these HMGTs reveals interesting relationships between gene function, phylogenetic distance, and frequency of multi-gene transfer. Among other insights, we find that (i) the observed relative frequency of HMGT increases as divergence between genomes increases, (ii) HMGTs often have conserved gene functions, and (iii) rare genes are frequently acquired through HMGT. We also analyze in detail HMGTs involving the zonula occludens toxin and type III secretion systems. By enabling the systematic inference of HMGTs on a large scale, HoMer will facilitate a more accurate and more complete understanding of HGT and microbial evolution.

Download Full-text

Evidence for a Type III Secretion System in Aeromonas salmonicida subsp. salmonicida

Journal of Bacteriology ◽

10.1128/jb.184.21.5966-5970.2002 ◽

2002 ◽

Vol 184 (21) ◽

pp. 5966-5970 ◽

Cited By ~ 74

Author(s):

Sarah E. Burr ◽

Katja Stuber ◽

Thomas Wahli ◽

Joachim Frey

Keyword(s):

Type Iii Secretion ◽

Type Strain ◽

Wild Type Strain ◽

Type Iii Secretion System ◽

Secretion System ◽

Aeromonas Salmonicida ◽

Broad Host Range ◽

Wild Type ◽

Secretion Systems ◽

Type Iii

ABSTRACT Aeromonas salmonicida subsp. salmonicida, the etiological agent of furunculosis, is an important fish pathogen. We have screened this bacterium with a broad-host-range probe directed against yscV, the gene that encodes the archetype of a highly conserved family of inner membrane proteins found in every known type III secretion system. This has led to the identification of seven open reading frames that encode homologues to proteins functioning within the type III secretion systems of Yersinia species. Six of these proteins are encoded by genes comprising a virA operon. The A. salmonicida subsp. salmonicida yscV homologue, ascV, was inactivated by marker replacement mutagenesis and used to generate an isogenic ascV mutant. Comparison of the extracellular protein profiles from the ascV mutant and the wild-type strain indicates that A. salmonicida subsp. salmonicida secretes proteins via a type III secretion system. The recently identified ADP-ribosylating toxin AexT was identified as one such protein. Finally, we have compared the toxicities of the wild-type A. salmonicida subsp. salmonicida strain and the ascV mutant against RTG-2 rainbow trout gonad cells. While infection with the wild-type strain results in significant morphological changes, including cell rounding, infection with the ascV mutant has no toxic effect, indicating that the type III secretion system we have identified plays an important role in the virulence of this pathogen.

Download Full-text