Comparative Genomic Analysis of Enterococci across Sectors of the One Health Continuum.

Enterococci are Gram-positive bacteria that can be isolated from a variety of environments including soil, water, plants, and the intestinal tract of humans and animals. Although they are considered commensals in humans, spp. are important opportunistic pathogens.

Resistance determinants and their genetic context in enterobacteria from a longitudinal study of pigs reared under various husbandry conditions.

Pigs are major reservoirs of resistant that can reach humans through consumption of contaminated meat or vegetables grown in manure-fertilized soil. Samples were collected from sows during lactation and their piglets at five time points spanning the production cycle. Cefotaxime-resistant bacteria were quantified and isolated from feed, feces, manures and carcasses of pigs reared with penicillin-using or antibiotic-free husbandries.

A λ Cro-Like Repressor Is Essential for the Induction of Conjugative Transfer of SXT/R391 Elements in Response to DNA Damage.

Unlabelled: Integrative and conjugative elements (ICEs) of the SXT/R391 family are the main contributors to acquired multidrug resistance in the seventh pandemic lineage of Vibrio cholerae, the etiological agent of the diarrheal disease cholera. Conjugative transfer of SXT/R391 ICEs is triggered by antibiotics and agents promoting DNA damage through RecA-dependent autoproteolysis of SetR, an ICE-encoded λ CI-like repressor. Here, we describe the role of CroS, a distant λ Cro homolog, as a key component contributing to the regulation of expression of the activator SetCD that orchestrates the expression of the conjugative transfer genes.

Electrophoretic Mobility Shift Assay Using Radiolabeled DNA Probes.

Electrophoretic mobility shift assays (EMSA) have proven their usefulness for studying interactions between biological molecules. In the present protocol, a purified protein of interest is mixed with a 5'-end radiolabeled DNA probe. The bound complexes are separated by electrophoretic migration through a polyacrylamide gel and detected with a phosphorimager.

The extended regulatory networks of SXT/R391 integrative and conjugative elements and IncA/C conjugative plasmids.

Nowadays, healthcare systems are challenged by a major worldwide drug resistance crisis caused by the massive and rapid dissemination of antibiotic resistance genes and associated emergence of multidrug resistant pathogenic bacteria, in both clinical and environmental settings. Conjugation is the main driving force of gene transfer among microorganisms. This mechanism of horizontal gene transfer mediates the translocation of large DNA fragments between two bacterial cells in direct contact.

A diaminopimelic acid auxotrophic Escherichia coli donor provides improved counterselection following intergeneric conjugation with actinomycetes.

Considering the medical, biotechnological, and economical importance of actinobacteria, there is a continuous need to improve the tools for genetic engineering of a broad range of these microorganisms. Intergeneric conjugation has proven to be a valuable yet imperfect tool for this purpose. The natural resistance of many actinomycetes to nalidixic acid (Nal) is generally exploited to eliminate the sensitive Escherichia coli donor strain following conjugation.

Transfer activation of SXT/R391 integrative and conjugative elements: unraveling the SetCD regulon.

Integrative and conjugative elements (ICEs) of the SXT/R391 family have been recognized as key drivers of antibiotic resistance dissemination in the seventh-pandemic lineage of Vibrio cholerae. SXT/R391 ICEs propagate by conjugation and integrate site-specifically into the chromosome of a wide range of environmental and clinical Gammaproteobacteria. SXT/R391 ICEs bear setC and setD, two conserved genes coding for a transcriptional activator complex that is essential for activation of conjugative transfer.

DNA-damaging agents induce the RecA-independent homologous recombination functions of integrating conjugative elements of the SXT/R391 family.

Integrating conjugative elements (ICEs) of the SXT/R391 family are major contributors to the spread of antibiotic resistance genes. These elements also catalyze their own diversity by promoting inter-ICE recombination through the action of the RecA-independent homologous recombination system that they encode. Here, we report that expression of this recombination system, which consists of the single-stranded DNA annealing protein Bet and the exonuclease Exo, is induced by DNA-damaging agents via ICE-encoded transcriptional regulators.

Dynamics of the SetCD-regulated integration and excision of genomic islands mobilized by integrating conjugative elements of the SXT/R391 family.

Mobilizable genomic islands (MGIs) are small genomic islands that are mobilizable by SXT/R391 integrating conjugative elements (ICEs) due to similar origins of transfer. Their site-specific integration and excision are catalyzed by the integrase that they encode, but their conjugative transfer entirely depends upon the conjugative machinery of SXT/R391 ICEs. In this study, we report the mechanisms that control the excision and integration processes of MGIs.

Chitosanase from Streptomyces coelicolor A3(2): biochemical properties and role in protection against antibacterial effect of chitosan.

Chitosan, an N-deacetylated derivative of chitin, has attracted much attention as an antimicrobial agent against fungi, bacteria, and viruses. Chitosanases, the glycoside hydrolases responsible for chitosan depolymerisation, are intensively studied as tools for biotechnological transformation of chitosan. The chitosanase CsnA (SCO0677) from Streptomyces coelicolor A3(2) was purified and characterized.