Draft Genome Sequence of an Escherichia coli O157:H43 Strain Isolated from Cattle

Here we report the draft genome sequence of an Escherichia coli O157:H43 strain, designated T22, with an atypical virulence gene profile and isolated from healthy cattle. T22 produces cytolethal distending toxin V (CDT-V) and belongs to phylogenetic group B1 and sequence type 155 (ST155)

The long polar fimbriae (lpf) operon and its flanking regions in bovine Escherichia coli O157:H43 and STEC O136:H12 strains

Long polar fimbriae (Lpf) are intestinal adhesins and important virulence factors of pathogenic Escherichia coli strains. We cloned and sequenced the lpf2-1 operon (lpf2ABCD) and its flanking regions of an intimin- and Shiga toxin-negative E. coli O157:H43 strain from bovine origin, and also sequenced the lpf2-1 operon of 6 additional atypical O157 bovine Escherichia coli strains of various serotypes Nucleotide sequence comparison of these lpf operons showed sequence conservation as they contain only four polymorphic nucleotide positions. Investigation of these O157 strains as well as 13 Escherichia coli Reference Collection (ECOR) strains carrying the lpf2-1 allele revealed high degree of sequence conservation in the lpf2 flanking regions. The lpf2-1 allele is also present in a bovine Shiga toxin-producing E. coli STEC O136:H12 strain and in vitro adherence assays revealed that the absence of lpf2-1 in this strain did not affect its host cell-binding properties. Our data indicate that lpf2 loci is highly conserved in E. coli isolates, but its role in adherence might be masked by other uncharacterized adhesins

Cytolethal distending toxin producing Escherichia coli O157:H43 strain T22 represents a novel evolutionary lineage within the O157 serogroup

Enterohemorrhagic Escherichia coli (EHEC) O157:H7/NMstrains are significant foodborne pathogens intensively studied, while other sero- and pathotypes of the O157 serogroup only began to receive more attention. Here we report the first genome sequence of a cytolethal distending toxin (CDT-V) producing E. coli O157:H43 strain (T22) isolated from cattle. The genome consists of a 4.9 Mb chromosome assembled into three contigs and one plasmid of 82.4 kb. Comparative genomic investigations conducted with the core genomes of representative E. coli strains in GenBank (n = 62) confirmed the separation of T22 from the EHEC and enteropathogenic (EPEC) O157 lineages. Gene content based pangenome analysis revealed as many as 261 T22-specific coding sequences without orthologs in EDL933 EHEC O157 prototypic and two phylogenetically related commensal E. coli strains. The genome sequence revealed 10 prophage-like regions which harbor several virulence-associated genes including cdt and heat-labile enterotoxin (LT-II) encoding operons. Our results indicate that the evolutionary path of T22 is largely independent from that of EHEC and EPEC O157:H7/NM strains. Thus, the CDT-producing T22 E. coli O157:H43 strain represents a unique lineage of E. coli O157

Methylene Blue Bridges the Inhibition and Produces Unusual Respiratory Changes in Complex III-Inhibited Mitochondria. Studies on Rats, Mice and Guinea Pigs

Methylene blue (MB) is used in human therapy in various pathological conditions. Its effects in neurodegenerative disease models are promising. MB acts on multiple cellular targets and mechanisms, but many of its potential beneficial effects are ascribed to be mitochondrial. Ac-cording to the “alternative electron transport” hypothesis, MB is capable of donating electrons to cytochrome c bypassing complex I and III. As a consequence of this, the deleterious effects of the inhibitors of complex I and III can be ameliorated by MB. Recently, the beneficial effects of MB ex-erted on complex III-inhibited mitochondria were debated. In the present contribution, several pieces of evidence are provided towards that MB is able to reduce cytochrome c and improve bio-energetic parameters, like respiration and membrane potential, in mitochondria treated with com-plex III inhibitors, either antimycin or myxothiazol. These conclusions were drawn from meas-urements for mitochondrial oxygen consumption, membrane potential, NAD(P)H steady state, MB uptake and MB-cytochrome c oxidoreduction. In the presence of MB and complex III inhibitors, unusual respiratory reactions, like decreased oxygen consumption as a response to ADP addition as well as stimulation of respiration upon administration of inhibitors of ATP synthase or ANT, were observed. Qualitatively identical results were obtained in three rodent species. The actual metabolic status of mitochondria is well reflected in the distribution of MB amongst various com-partments of this organelle

R18C is a new viable P2-like bacteriophage of rabbit origin infecting Citrobacter rodentium and Shigella sonnei strains

Here we report a novel virulent P2-like bacteriophage R18C isolated from rabbit faeces, which, additionally to Escherichia coli K-12 strains, was able to propagate on Citrobacter rodentium strain ICC169 and a range of Shigella sonnei with high efficiency of plating (EOP). It represents the first lytic bacteriophage originated from rabbit and the first infective P2-like phage of animal origin. In the three characteristic moron-harboring regions of P2-like phages, R18C harbors genes with unknown function, which have been only found in cryptic P2-like prophages up to date

A novel transducible chimeric phage from Escherichia coli O157:H7 Sakai strain encoding Stx1 production.

Shiga toxin-producing Escherichia coli (STEC), and especially enterohaemorrhagic E. coli (EHEC) are important, highly virulent zoonotic and food-borne pathogens. The genes encoding their key virulence factors, the Shiga toxins, are distributed by converting bacteriophages, the Stx phages. In this study we isolated a new type of inducible Stx phage carrying the stx1 gene cluster from the prototypic EHEC O157:H7 Sakai strain. The phage showed Podoviridae morphology, and was capable of converting the E. coli K-12 MG1655 strain to Shiga toxin-producing phenotype. The majority of the phage genes originate from the stx2-encoding Sakai prophage Sp5, with major rearrangements in its genome. Beside certain minor recombinations, the genomic region originally containing the stx2 genes in Sp5 was replaced by a region containing six open reading frames from prophage Sp15 including stx1 genes. The rearranged genome, together with the carriage of stx1 genes, the morphology and the capability of lysogenic conversion represent a new type of recombinant Stx1 converting phage from the Sakai strain

Comparative Genomic and Phylogenetic Analysis of a Shiga Toxin Producing Shigella sonnei (STSS) Strain

Shigella strains are important agents of bacillary dysentery, and in recent years Shigella sonnei has emerged as the leading cause of shigellosis in industrialized and rapidly developing countries. More recently, several S. sonnei and Shigella flexneri strains producing Shiga toxin (Stx) have been reported from sporadic cases and from an outbreak in America. In the present study we aimed to shed light on the evolution of a recently identified Shiga toxin producing S. sonnei (STSS) isolated in Europe. Here we report the first completely assembled whole genome sequence of a multidrug resistant (MDR) Stx-producing S. sonnei (STSS) clinical strain and reveal its phylogenetic relations. STSS 75/02 proved to be resistant to ampicillin, streptomycin, tetracycline, chloramphenicol, thrimetoprim, and sulfomethoxazol. The genome of STSS 75/02 contains a 4,891,717 nt chromosome and seven plasmids including the 214 kb invasion plasmid (pInv) harboring type III secretion system genes and associated effectors. The chromosome harbors 23 prophage regions including the Stx1 converting prophage. The genome carries all virulence determinants necessary for an enteroinvasive lifestyle, as well as the Stx1 encoding gene cluster within an earlier described inducible converting prophage. In silico SNP genotyping of the assembled genome as well as 438 complete or draft S. sonnei genomes downloaded from NCBI GenBank revealed that S. sonnei 75/02 belongs to the more recently diverged global MDR lineage (IIIc). Targeted screening of 1131 next-generation sequencing projects taken from NCBI Short Read Archive of confirms that only a few S. sonnei isolates are Stx positive. Our results suggest that the acquisition of Stx phages could have occurred in different environments as independent events and that multiple horizontal transfers are responsible for the appearance of Stx phages in S. sonnei strains

Methylene Blue Bridges the Inhibition and Produces Unusual Respiratory Changes in Complex III-Inhibited Mitochondria. Studies on Rats, Mice and Guinea Pigs

Methylene blue (MB) is used in human therapy in various pathological conditions. Its effects in neurodegenerative disease models are promising. MB acts on multiple cellular targets and mechanisms, but many of its potential beneficial effects are ascribed to be mitochondrial. According to the “alternative electron transport” hypothesis, MB is capable of donating electrons to cytochrome c bypassing complex I and III. As a consequence of this, the deleterious effects of the inhibitors of complex I and III can be ameliorated by MB. Recently, the beneficial effects of MB exerted on complex III-inhibited mitochondria were debated. In the present contribution, several pieces of evidence are provided towards that MB is able to reduce cytochrome c and improve bioenergetic parameters, like respiration and membrane potential, in mitochondria treated with complex III inhibitors, either antimycin or myxothiazol. These conclusions were drawn from measurements for mitochondrial oxygen consumption, membrane potential, NAD(P)H steady state, MB uptake and MB-cytochrome c oxidoreduction. In the presence of MB and complex III inhibitors, unusual respiratory reactions, like decreased oxygen consumption as a response to ADP addition as well as stimulation of respiration upon administration of inhibitors of ATP synthase or ANT, were observed. Qualitatively identical results were obtained in three rodent species. The actual metabolic status of mitochondria is well reflected in the distribution of MB amongst various compartments of this organelle

Comparative analysis of the Shiga toxin converting bacteriophage first detected in Shigella sonnei

Here we report the first complete nucleotide sequence of a Shiga toxin (Stx) converting phage from a Shigella sonnei clinical isolate that harbors stx1 operon, first identified in the chromosome of Shigella dysenteriae type 1. The phage named Shigella phage 75/02 Stx displayed Podoviridae morphology. It proved to be transferable to Escherichia coli K-12 strains, and cytotoxicity of the lysogenized strains was demonstrated in Vero cell cultures. Genomic analysis revealed that the prophage genome is circular and its size is 60,875 nt that corresponds to 76 ORFs. The genome of Shigella phage 75/02 Stx shows a great degree of mosaic structure and its architecture is related to lambdoid phages. All the deduced proteins, including the 37 hypothetical proteins showed significant homologies to Stx phage proteins present in databases. The phage uniformly inserted into the ynfG oxidoreductase gene framed by phage integrase and antirepressor genes in parental S. sonnei and in the three lysogenized K-12 strains C600, DH5α and MG1655. The Stx1 prophage proved to be stable in its bacterial hosts and remained inducible