Mouse lung infection model to assess Rhodococcus equi virulence and vaccine protection

González-Iglesias, Patricia; Scortti, Mariela; MacArthur, Iain; Hapeshi, Alexia; Rodríguez, Héctor; Prescott, John F.; Vázquez‐Boland, José A.

doi:10.1016/j.vetmic.2014.03.026

Cited by 17 publications

(29 citation statements)

References 46 publications

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“…R. equi PAM1571 is a prototypic traA ϩ vapAB-negative bovine strain (34) isolated from a heifer's mediastinal lymph node with pyogranulomatous lesions (kindly provided by F. Quigley, Central Veterinary Research Laboratory, Ireland) (12). Its plasmid-cured derivative, PAM1571 Ϫ , was obtained by subjecting wildtype bacteria to an electroporation pulse of 12.5 kV/cm at 1,000 ⍀ and 25 F (GenePulser Xcell; Bio-Rad) followed by six cycles of plating and single-colony subculturing in liquid medium at 37°C (27). These two strains are henceforth designated 1571 and 1571 Ϫ , respectively.…”

Section: Methodsmentioning

confidence: 99%

“…R. equi 103S is the reference genome strain, a low-passage clone of equine clinical isolate 103 ϩ used in different laboratories (24). Its isogenic 103S⌬vapA and plasmid-cured 103S Ϫ derivatives were described previously (27). The presence of the virulence plasmid was routinely checked in all strains by PCR using suitable oligonucleotide primers (see Table S1 in the supplemental material).…”

Section: Methodsmentioning

confidence: 99%

“…Total DNA extraction and purification from R. equi, PCR techniques, DNA fragment purification and electrophoresis, recombinant DNA techniques, and plasmid purification and electroporation were performed as previously described (27,34,36). For pulsed-field gel electrophoresis (PFGE), plugs were formed by embedding R. equi cells from 1-ml aliquots of stationary (24-h) BHI cultures in melted 1% agarose Tris-EDTA (TE).…”

Section: Methodsmentioning

confidence: 99%

“…Mouse intranasal and intravenous (i.v.) infections and lung competitive virulence assays were performed as previously described (27). The relative proportions of competing bacteria were calculated by analyzing at least 40 random colonies from the plated organ homogenate by PCR using suitable oligonucleotide primers (see Table S1 in the supplemental material).…”

Section: Smentioning

confidence: 99%

“…One of them, VapA, a 19-kDa secreted protein, is essential for virulence. A single vapA gene deletion causes strong attenuation comparable to that caused by loss of the plasmid, with an inability to both proliferate in macrophages and survive in vivo in mice (26,27).…”

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confidence: 99%

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An Invertron-Like Linear Plasmid Mediates Intracellular Survival and Virulence in Bovine Isolates of Rhodococcus equi

et al. 2015

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dWe report a novel host-associated virulence plasmid in Rhodococcus equi, pVAPN, carried by bovine isolates of this facultative intracellular pathogenic actinomycete. Surprisingly, pVAPN is a 120-kb invertron-like linear replicon unrelated to the circular virulence plasmids associated with equine (pVAPA) and porcine (pVAPB variant) R. equi isolates. pVAPN is similar to the linear plasmid pNSL1 from Rhodococcus sp. NS1 and harbors six new vap multigene family members (vapN to vapS) in a vap pathogenicity locus presumably acquired via en bloc mobilization from a direct predecessor of equine pVAPA. Loss of pVAPN rendered R. equi avirulent in macrophages and mice. Mating experiments using an in vivo transconjugant selection strategy demonstrated that pVAPN transfer is sufficient to confer virulence to a plasmid-cured R. equi recipient. Phylogenetic analyses assigned the vap multigene family complement from pVAPN, pVAPA, and pVAPB to seven monophyletic clades, each containing plasmid type-specific allelic variants of a precursor vap gene carried by the nearest vap island ancestor. Deletion of vapN, the predicted "bovine-type" allelic counterpart of vapA, essential for virulence in pVAPA, abrogated pVAPN-mediated intramacrophage proliferation and virulence in mice. Our findings support a model in which R. equi virulence is conferred by host-adapted plasmids. Their central role is mediating intracellular proliferation in macrophages, promoted by a key vap determinant present in the common ancestor of the plasmid-specific vap islands, with host tropism as a secondary trait selected during coevolution with specific animal species.

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Smentioning

confidence: 99%

mentioning

confidence: 99%

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An Invertron-Like Linear Plasmid Mediates Intracellular Survival and Virulence in Bovine Isolates of Rhodococcus equi

et al. 2015

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The Rhodococcus equi virulence protein VapA disrupts endolysosome function and stimulates lysosome biogenesis

et al. 2016

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Rhodococcus equi (R. equi) is an important pulmonary pathogen in foals that often leads to the death of the horse. The bacterium harbors a virulence plasmid that encodes numerous virulence‐associated proteins (Vaps) including VapA that is essential for intracellular survival inside macrophages. However, little is known about the precise function of VapA. Here, we demonstrate that VapA causes perturbation to late endocytic organelles with swollen endolysosome organelles having reduced Cathepsin B activity and an accumulation of LBPA, LC3 and Rab7. The data are indicative of a loss of endolysosomal function, which leads cells to upregulate lysosome biogenesis to compensate for the loss of functional endolysosomes. Although there is a high degree of homology of the core region of VapA to other Vap proteins, only the highly conserved core region of VapA, and not VapD of VapG, gives the observed effects on endolysosomes. This is the first demonstration of how VapA works and implies that VapA aids R. equi survival by reducing the impact of lysosomes on phagocytosed bacteria.

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Rhodococcus equi–What is New This Decade?

Sanz¹

2023

Veterinary Clinics of North America: Equine Practice

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Mouse lung infection model to assess Rhodococcus equi virulence and vaccine protection

Cited by 17 publications

References 46 publications

An Invertron-Like Linear Plasmid Mediates Intracellular Survival and Virulence in Bovine Isolates of Rhodococcus equi

An Invertron-Like Linear Plasmid Mediates Intracellular Survival and Virulence in Bovine Isolates of Rhodococcus equi

The Rhodococcus equi virulence protein VapA disrupts endolysosome function and stimulates lysosome biogenesis

Rhodococcus equi–What is New This Decade?

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