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Rhodococcus equi is a soilborne aerobic actinomycete bacterium that infects animals and humans. Human infections are opportunistic, zoonotic in origin, and considered to be linked to exposure to farm environments [...]

Rhodococcus equi is a soilbome facultative intracellular actinobacterium that causes pyogranulomatous infections in multiple animal species, including humans. Rhodococcal infection is particularly severe in young foals and immunocompromised persons, in [...]

The pathogenic actinomycete Rhodococcus equi harbors different types of virulence plasmids associated with specific nonhuman hosts. We determined the complete DNA sequence of a [vapB.sup.+] plasmid, typically associated with pig isolates, and compared it with that of the horse-specific [vapA.sup.+] plasmid type. pVAPB1593, a circular 79,251-bp element, had the same housekeeping backbone as the [vapA.sup.+] plasmid but differed over an [approximately equal to] 22-kb region. This variable region encompassed the vap pathogenicity island (PAI), was clearly subject to selective pressures different from those affecting the backbone, and showed major genetic rearrangements involving the vap genes. The pVAPB1593 PAI harbored five different vap genes (vapB and vapJ to -M, with vapK present in two copies), which encoded products differing by 24 to 84% in amino acid sequence from the six full-length [vapA.sup.+] plasmid-encoded Vap proteins, consistent with a role for the specific vap gene complement in R. equi host tropism. Sequence analyses, including interpolated variable-order motifs for detection of alien DNA and reconstruction of Vap family phylogenetic relationships, suggested that the vap PAI was acquired by an ancestor plasmid via lateral gene transfer, subsequently evolving by vap gene duplication and sequence diversification to give different (host-adapted) plasmids. The R. equi virulence plasmids belong to a new family of actinobacterial circular replicons characterized by an ancient conjugative backbone and a horizontally acquired niche-adaptive plasticity region.

Little is known about the iron acquisition systems of the soilborne facultative intraceilular pathogen Rhodococcus equi. We previously reported that expression of iupABC, encoding a putative siderophore ABC transporter system, is iron regulated and required for growth at low iron concentrations. Here we show that disruption of iupA leads to the concomitant accumulation of catecholates and a chromophore with absorption maxima at 341 and 528 nm during growth under iron-replete conditions. In contrast, the wild-type strain produces these compounds only in iron-depleted medium. Disruption of iupU and iupS, encoding nonribosomal peptide synthetases, prevented growth of the corresponding R. equi SID1 and SID3 mutants at low iron concentrations. However, only R. equi SID3 did not produce the chromophore produced by the wild-type strain during growth at low iron concentrations. The phenotype of R. equi SID3, but not that of R. equi SID1, could be rescued by coculture with the wild type, allowing growth at low iron concentrations. This strongly suggests that the product of the iupS gene is responsible for the synthesis of a diffusible compound required for growth at low iron concentrations. Transcription of iupU was constitutive, but that of iupS was iron regulated, with an induction of 3 orders of magnitude during growth in iron-depleted compared to iron-replete medium. Neither mutant was attenuated in vivo in a mouse infection model, indicating that the iupU- and iupS-encoded iron acquisition systems are primarily involved in iron uptake during saprophytic life.

The development and optimization of a novel quantitative real-time (Q)-PCR assay for Listeria monocytogenes is reported based on the simultaneous detection of hly gene target sequences, which provides high specificity, sensitivity and quantifiability, and an internal amplification control (IAC) sequence for the assessment of PCR inhibition. This Q-PCR assay for Listeria monocytogenes is a simple and robust tool that facilitates the identification of false negatives or underestimations of contamination loads due to PCR failure.

Efficient replication in vivo is essential for a microparasite to colonize its host and the understanding of the molecular mechanisms by which microbial pathogens grow within host tissues can lead to the discovery of novel therapies to treat infection. Here we present evidence that the foodborne bacterial pathogen Listeria monocytogenes, a facultative intracellular parasite, exploits hexose phosphates (HP) from the host cell as a source of carbon and energy to fuel fast intracellular growth. HP uptake is mediated by Hpt, a bacterial homolog of the mammalian translocase that transports glucose-6-phosphate from the cytosol into the endoplasmic reticulum in the final step of gluconeogenesis and glycogenolysis. Expression of the Hpt permease is tightly controlled by the central virulence regulator PrfA, which upon entry into host cells induces a set of virulence factors required for listerial intracellular parasitism. Loss of Hpt resulted in impaired listerial intracytosolic proliferation and attenuated virulence in mice. Hpt is the first virulence factor to be identified as specifically involved in the replication phase of a facultative intracellular pathogen. It is also a clear example of how adaptation to intracellular parasitism by microbial pathogens involves mimicry of physiological mechanisms of their eukaryotic host cells.

Research reveals that the choE monocistron gene from Rhodococcus equi encodes a membrane-damaging toxin and a virulence factor, cholesterol oxidase. Data indicate that ChoE is homologous to other bacterial cholesterol oxidases and functions as a cytolytic factor in R. equi.

The effect of carbohydrates on virulence gene expression was investigated in a panel of wild-type isolates of Listeria monocytogenes by employing the PrfA-dependent phospholipase C gene plcB as a reporter. Results showed that the bvr locus encodes a beta-glucoside-specific sensor that mediates virulence gene repression upon detection of cellobiose and salicin. Bvr is the first sensory system found in L. monocytogenes that is implicated in environmental regulation of virulence genes.

Most Listeria monocytogenes virulence genes are positively controlled by the PrfA protein. Mutants of the prfA (prfA*), which as a result of single amino acid substitution in PrfA, Gly145Ser, constitutively overexpress prfA and the genes for the PrfA virulence regulon, have been previously characterized. It is demonstrated that approximately 10 times more PrfA protein is produced in a prfA* strain than in the wild-type.

A study was conducted on glucose-1-phosphate (G-1-P) and its utilization by Listeria monocytogenes under the tight positive control of PrfA, the central regulator of listerial virulence. The findings suggest that G-1-P could play a key role as a growth substrate for intracellular Listeria. Moreover, it was found that the catabolism of G-1-P is coordinately expressed with virulence genes.

A suicide conjugative vector, pLOF/km, carrying a mini-Tn10 delivery system, aids in developing a transposon mutagenesis system, which is functional in the gram negative swine pathogen Actinobacillus pleuropneumoniae. In the genome of A. pleuropneumoniae, insertional disruptions are produced with the help of the transposon mutagenesis system that is random, single and stable at a workable frequency. The transposon mutagenesis system is better than other methods, and will aid in further gentic research of A. pleuropneumoniae.

The genus Listeria contains 2 pathogenic species, L. monocytogenes and L. ivanovii (1). They both invade host cells, replicate in the cytosol after phagosomal escape, and spread from cell to [...]

Author(s): Mariela Scortti [1, 2, 3, 4]; Lizeth Lacharme-Lora [1, 3, 4]; Martin Wagner [2, 4, 5]; Isabel Chico-Calero [2]; Patrizia Losito [1]; Jose A Vazquez-Boland (corresponding author) [1, 2, [...]

The article informs about the emergence of multidrug-resistant Rhodococcus equi (MDR-RE), with isolates demonstrating resistance to antibiotics including macrolides and rifampicin. Topic includes the global dissemination risk of MDR-RE, originating from a specific lineage likely traced back to Kentucky and circulating in equine farms in New York state, emphasizing the need for clinical efficacy studies to find alternative treatments for MDR-RE infections in foals.

The usefulness of hly and iap genes as targets for the specific quantitative detection of L. monocytogenes by RTi-PCR is evaluated. The Amplifluor and TaqMan technologies are used in the experiment.

We report a comparative study of 29 representative genomes of the animal pathogen Rhodococcus equi. The analyses showed that R. equi is genetically homogeneous and clonal, with a large core genome accounting for ≈80% of an isolates' gene content. An open pangenome, even distribution of accessory genes among the isolates, and absence of significant core-genome recombination, indicated that gene gain/loss is a main driver of R. equi genome evolution. Traits previously predicted to be important in R. equi physiology, virulence and niche adaptation were part of the core genome. This included the lack of a phosphoenolpyruvate:carbohydrate transport system (PTS), unique among the rhodococci except for the closely related Rhodococcus defluvii, reflecting selective PTS gene loss in the R. equi-R. defluvii sublineage. Thought to be asaccharolytic, rbsCB and glcP non-PTS sugar permease homologues were identified in the core genome and, albeit inefficiently, R. equi utilized their putative substrates, ribose and (irregularly) glucose. There was no correlation between R. equi whole-genome phylogeny and host or geographical source, with evidence of global spread of genomovars. The distribution of host-associated virulence plasmid types was consistent with the exchange of the plasmids (and corresponding host shifts) across the R. equi population, and human infection being zoonotic. Phylogenomic analyses demonstrated that R. equi occupies a central position in the Rhodococcus phylogeny, not supporting the recently proposed transfer of the species to a new genus. [ABSTRACT FROM AUTHOR]

Peptidoglycan recognition proteins are a family of evolutionary conserved proteins that play a basic role in the innate immunity of insects, but their role in the immunity of mammals remains unclear. To elucidate its functions, a mouse member of the peptidoglycan recognition proteins family, TagL, was stably expressed in colon adenocarcinoma HT29 cells, and its effect on the invasion and intracellular growth of the enteroinvasive pathogenic bacterium Listeria monocytogenes was assessed. The expression of TagL substantially impaired bacterial invasion and early intracellular growth. The observed effects were partly caused by a loss of viability by intraphagosomal bacteria. Efficient phagosome escaping but not efficient invasion helped bacteria to overplay TagL. [ABSTRACT FROM AUTHOR]

Identifies an internalin locus of Listeria ivanovii. Etiological agents of foodborne infection called listeriosis; Involvement of several virulence factors in intracellular survival and cell-to-cell spread; Bacterial strains; Construction of mutant strains; Cell culture assays.