Michal LetekUniversidad de León | UNILEON · Department of Biochemistry and Molecular Biology
Michal Letek
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Introduction
I have an interest in studying actinobacterial virulence to identify novel ways to block the pathogenesis of these bacteria. I functionally analyzed the genome of an intracellular pathogen, Rhodococcus equi. The reconstruction of the regulatory network of this pathogen led to the discovery of novel virulence factors used by R. equi to survive inside macrophages. I am now studying its redox biology during cell infection.
My research work is also focused on the identification and characterization of the host molecular factors that are involved in the intracellular survival and proliferation of Staphylococcus aureus, a major human pathogen. This could be crucial to design novel therapeutic strategies to control MRSA and to identify the major determinants of susceptibility to S. aureus.
Skills and Expertise
Publications
Publications (84)
The development of RNA-based anti-infectives has gained interest with the successful application of mRNA-based vaccines. Small RNAs are molecules of RNA of <200 nucleotides in length that may control the expression of specific genes. Small RNAs include small interference RNAs (siRNAs), Piwi-interacting RNAs (piRNAs), or microRNAs (miRNAs). Notably,...
Antibiotherapy is the main therapeutic strategy in the fight against bacterial pathogens. However, the misuse of antimicrobials has led to the appearance of antimicrobial-resistant strains. The rate at which we isolate multidrug-resistant bacteria is now much faster than the discovery rate of new antimicrobials. Therefore, the repurposing of approv...
Every year, we face infectious outbreaks produced by harmful microorganisms commonly called superbugs. Often, there is not enough time to find new treatments to cure infected patients. On average, it takes a decade to develop a promising new drug to the point where it can be used on patients! Also, many of the compounds that we identify in the labo...
Staphylococcal infections are a widespread cause of disease in humans. In particular, S. aureus is a major causative agent of infection in clinical medicine. In addition, these bacteria can produce a high number of staphylococcal enterotoxins (SE) that may cause food intoxications. Apart from S. aureus, many coagulase-negative Staphylococcus spp. c...
Rhodococcus equi is an intracellular veterinary pathogen that is becoming resistant to current antibiotherapy. Genes involved in preserving redox homeostasis could be promising targets for the development of novel anti-infectives. Here, we studied the role of an extracellular thioredoxin (Etrx3/REQ_13520) in the resistance to phagocytosis. An etrx3...
Probiotics have been shown to bind to host receptors, which are important for pathogen adhesion and induce the host’s production of defence factors. They can activate the goblet-cell-derived production of mucins, a major component of the mucus layer and a physical barrier participating in limiting the proximity of microorganisms to the epithelial l...
The genus Staphylococcus encompasses many species that may be pathogenic to both humans and farm animals. These bacteria have the potential to acquire multiple resistant traits to the antimicrobials currently used in the veterinary or medical settings. These pathogens may commonly cause zoonoses, and the infections they cause are becoming difficult...
Tuberculosis is the leading cause of death, worldwide, due to a bacterial pathogen. This respiratory disease is caused by the intracellular pathogen Mycobacterium tuberculosis and produces 1.5 million deaths every year. The incidence of tuberculosis has decreased during the last decade, but the emergence of MultiDrug-Resistant (MDR-TB) and Extensiv...
Antimicrobial resistance is becoming one of the most important human health issues. Accordingly, the research focused on finding new antibiotherapeutic strategies is again becoming a priority for governments and major funding bodies. The development of treatments based on the generation of oxidative stress with the aim to disrupt the redox defenses...
Rhodococcus equi is a facultative intracellular pathogen that causes infections in foals and
many other animals such as pigs, cattle, sheep, and goats. Antibiotic resistance is rapidly rising in horse
farms, which makes ine�ective current antibiotic treatments based on a combination of macrolides and
rifampicin. Therefore, new therapeutic strategie...
In 1928, Sir Alexander Fleming observed the bacterial-killing effects of penicillin in his laboratory in London. This was the first step in the discovery of one of the most important pillars of today’s medicine: the antibiotics. It took many years to find a way to produce penicillin in large amounts, and large-scale production did not start until 1...
Staphylococcus aureus is a facultative intracellular pathogen that invades and replicates within many types of human cells. S. aureus has shown to rapidly overcome traditional antibiotherapy by developing multidrug resistance. Furthermore, intracellular S. aureus is protected from the last-resort antibiotics—vancomycin, daptomycin, and linezolid—as...
Rhodococcus equi is a facultative intracellular pathogen that can survive within macrophages of a wide variety of hosts, including immunosuppressed humans. Current antibiotherapy is often ineffective, and novel therapeutic strategies are urgently needed to tackle infections caused by this pathogen. In this study, we identified three mycoredoxin-enc...
During patient colonization, Staphylococcus aureus is able to invade and proliferate within human cells to evade the immune system and last resort drugs such as vancomycin. Hijacking specific host molecular factors and/or pathways is necessary for pathogens to successfully establish an intracellular infection. In this study, we employed an unbiased...
As a facultative intracellular pathogen, Staphylococcus aureus is able to invade and proliferate within many types of mammalian cells. Intracellular bacterial replication relies on host nutrient supplies and, therefore, cell metabolism is closely bound to intracellular infection. Here, we investigated how S. aureus invasion affects the host membran...
Host-directed therapeutics are a promising anti-infective strategy against intracellular bacterial pathogens. Repurposing host-targeted drugs approved by the FDA in the US, the MHRA in the UK and/or regulatory equivalents in other countries, is particularly interesting because these drugs are commercially available, safe doses are documented and th...
Acetaminophen (APAP) is one of the most commonly used analgesics worldwide, and overdoses are associated with lactic acidosis, hepatocyte toxicity, and acute liver failure due to oxidative stress and mitochondrial dysfunction. Hepatoma cell lines typically lack the CYP450 activity to generate the reactive metabolite of APAP observed in vivo, but ar...
The infiltration of Th17 cells in tissues and organs during the development of many autoimmune diseases is considered a key step toward the establishment of chronic inflammation. Indeed, the localized and prolonged release of IL-17 in specific tissues has been associated with an increased severity of the inflammatory response that remains sustained...
Staphylococcus aureus escapes from immune recognition by invading a wide range of human cells. Once the pathogen becomes intracellular, the most important last resort antibiotics are not effective. Therefore, novel anti-infective therapies against intracellular S. aureus are urgently needed. Here, we have studied the physiological changes induced i...
Arsenic (As) is widespread in the environment and highly toxic. It has been released by volcanic and anthropogenic activities and causes serious health problems worldwide. To survive arsenic-rich environments, soil and saprophytic microorganisms have developed molecular detoxification mechanisms to survive arsenic-rich environments, mainly by the e...
Bacterial cell size and morphology are enormously diverse. The molecular
factors of morphogenesis are well understood in certain bacterial models and fairly
conserved throughout a broad spectrum of bacterial species, as follows. In most
bacteria, the tubulin-like FtsZ protein polymerizes at the mid cell , thereby generating
the scaffold of the bact...
The present clinical scenario is one of a growing number of immuno-compromised patients infected with a variety of fungal pathogens. AIDs, tuberculosis, immunosuppressive therapy, cancer chemotherapy or the use of broad-spectrum antibiotics contribute to the boost of such patient category. However, progress in the treatment of fungal infections has...
Selective autophagy underlies many of the important physiological roles that autophagy plays in multicellular organisms, but the mechanisms involved in cargo selection are poorly understood. Here we describe a molecular mechanism that can target conventional endosomes for autophagic degradation. We show that the human transmembrane protein TMEM59 c...
Adaptation to endoplasmic reticulum (ER) stress depends on the activation of the unfolded protein response (UPR) stress sensor inositol-requiring enzyme 1α (IRE1α), which functions as an endoribonuclease that splices the mRNA of the transcription factor XBP-1 (X-box-binding protein-1). Through a global proteomic approach we identified the BCL-2 fam...
Although bacteria are considered the simplest life forms, we are now slowly unraveling their cellular complexity. Surprisingly, not only do bacterial cells have a cytoskeleton but also the building blocks are not very different from the cytoskeleton that our own cells use to grow and divide. Nonetheless, despite important advances in our understand...
Layout file of expression network analysis with r = 0.95. Viewable with Biolayout Express 3D (http://www.biolayout.org/).
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Layout file of expression network analysis with r = 0.95 (nodes not belonging to plasmid gene-containing clusters have been removed). Viewable with Biolayout Express 3D (http://www.biolayout.org/).
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Layout file of expression network analysis with r = 0.85. Viewable with Biolayout Express 3D (http://www.biolayout.org/).
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Scatter plots of selected functional categories vs genome size (≥4 Mb) of R. equi 103S and 10 other representative Actinobacteria. Data were inferred using the Comprehensive Microbial Resource (http://cmr.jcvi.org/) and the available genomes (Data Release 23.0). See Table S13 for accession nos. Membrane-associated and secreted proteins, as determin...
Species-specific gene complements of R. equi 103S, R. jostii RHA1, N. farcinica IFM10152, and M. tuberculosis H37Rv. The Venn diagram shows the number of chromosomal CDSs shared within a particular relationship (in brackets those unique to that relationship) as determined by ortholog comparisons (reciprocal FASTA best hits). Below the name of each...
R. equi nutrition and metabolism. (A) Carbon source utilization. Growth assays of R. equi 103S in mineral medium (MM) [19] at 37°C. MM was supplemented (unless otherwise stated) with 20 mM of the indicated carbon sources and bacterial growth was monitored at OD600 every 30 min in a Fluostar Omega plate reader (BMG Labtech). Growth was detected only...
Circular diagram of the R. equi 103S genome (chromosome and virulence plasmid). Outer two rings, coding sequences in the forward and reverse strand colored according to functional class (see Figure S3). Left, R. equi 103S chromosome with ortholog comparison and horizontally acquired (HGT) islands. Ortholog plots from 13 actinobacterial genomes are...
Optimal growth pH of R. equi 103S. Phenotype MicroArray [15] output of the relevant wells of plate PM10. Incubation was for 48 h at 37°C in an OmniLog instrument with readings taken every 15 minutes. Data were analyzed with OmniLog PM software. Consensus phenotypes for at least two replicas were determined based on the area difference under the kin...
Genetic structure of the two large chromosomal HGT regions in R. equi 103S. The position of these regions on the chromosome is indicated in Figure S1. Functional categories of the genes are indicated in color code as in Figure S3. Alien Hunter [92] HGT hits are indicated as black bars in the center. HGT region 1 (positions 1,684,996-1,775,619, REQ1...
Examples of antibiotic resistance determinants located at the same chromosomal position in R. equi and two environmental Rhodococcus spp. Homologous resistance determinants indicated by yellow stripes in the ACT alignments.
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Statistics of horizontal gene acquisition (HGT) in actinobacterial chromosomes. HGT DNA was identified with the Alien Hunter program (http://www.sanger.ac.uk/Software/analysis/), which identifies horizontally acquired DNA by reliably capturing local compositional biases based on a variable-order motif distributions method [92]. The thick gray line...
Virulence plasmid-chromosome crosstalk. Gobal microarray expression analysis of R. equi 103S and an isogenic plasmid-cured derivative (103SP−) during exponential growth in LB medium (OD600 = 0.8) in the indicated conditions (part A of table, 30°C-pH 8.0 = vap PAI gene-downregulating conditions; part B of table, 37°C-pH 6.5 = vap PAI gene-activating...
Pairwise ACT alignments of rhodococcal chromosomes (R. equi 103S, R. jostii RHA1, R. opacus B4 and R. erythropolis PR4); see Figure 1A for interpretation. R. opacus has a large (7.25 Mb) linear chromosome like R. jostii (Table 1). The chromosome of R. erythropolis (6.52 Mb) is circular, as in R. equi. The four rhodococcal species sequenced to date...
Functional classification of R. equi 103S genome. According to the Ecocyc classification scheme [93]. (A) Functional categories of R. equi 103S genes. “Surface/extracellular proteins” includes products with a signal sequence and/or transmembrane domain not allocated to another main functional category (e.g. central metabolism, degradation of small...
Ranking of the ten most populated paralogous metabolic gene families of R. equi 103S, R. jostii RHA1, N. farcinica IFM10152, and M. tuberculosis H37Rv. Determined by BLASTCLUST analysis. In brackets, number of paralogs within the family.
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Potential virulence-associated genes of R. equi 103S identified by bioinformatic mining of the genome and homologs in other pathogenic and nonpathogenic Actinobacteria.
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Experimentally determined virulence-associated genes of M. tuberculosis and homologs in nonpathogenic Actinobacteria.
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Plasmid gene-containing coregulated clusters. Gene allocation defined by graph clustering of the transcription network shown in Figure 5A. (A) Plasmid backbone cluster. Shown for each gene, average pairwise comparison ratios of normalized microarray expression data from exponential cultures of R. equi 103S in LB medium (OD600 = 0.8) at 37°C relativ...
GenBank accession nos. of the genomes used in this study. R. erythropolis PR4 and R. opacus B4 genomes published online by NITE, the Japanese National Institute for Technology and Evaluation (http://www.nite.go.jp/index-e.html).
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Species-specific metabolic gene complements of R. equi 103S, R. jostii RHA1, N. farcinica IFM10152, and M. tuberculosis H37Rv. Determined by ortholog comparison (reciprocal FASTA best hits). As the functional categories used for the annotation of the four genomes were not directly comparable, we first extracted the metabolism-related CDSs manually,...
Phosphoenolpyruvate-sugar phosphotransferase system (PTS) components in a selection of actinobacterial genomes. Identified using motif search in Pfam database (Pfam motif identifiers indicated in footnotes).
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Minimal inhibitory concentrations (MIC) of R. equi 103S to various antibiotics. Determined by the broth microdilution method. The data are consistent with previously reported antimicrobial susceptibility studies of R. equi isolates [111]–[116].
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Software and databases used to annotate and analyze the R. equi 103S genome.
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Oligonucleotide primers used for mutant construction and complementation. SpeI, XbaI and EcoRV restriction sites used for the cloning of PCR products are underlined.
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Virulence-related loci of R. equi 103S. (A) PE/PPE locus and corresponding chromosomal regions in R. jostii RHA1, R. erythropolis PR4, N. farcinica IFM10152 and M. tuberculosis H37Rv. Arrows in ACT alignments indicate PE and PPE genes. The PE gene is of the “short” subclass (only a conserved N-terminal PE module of 99 to 102 residues); the PPE gene...
Network analysis of R. equi microarray expression data. (A) Detail of the network graph of Figure 5A showing the web of functional linkages (edges) between the vap PAI-coregulated cluster (red nodes) and direct neighbor clusters (green nodes, plasmid backbone cluster; other clusters represented in different colors; individual directly connected nod...
Chromosomal gene duplication and paralogous families in R. equi 103S and 19 other representative Actinobacteria. Paralogous families were identified by clustering of proteomes with BLASTClust (see Table S12).
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DNA mobility genes in R. equi 103S and environmental Rhodococcus spp genomes. Identified by keyword parsing of protein annotation; in brackets, genes associated with HGT regions. Plasmids from R. erythropolis PR4 published in [106].
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Putative DosR/DevR boxes and corresponding transcriptional units in R. equi 103S a. Identified with CLC Main Workbench (http://www.clcbio.com/) and the 20-bp consensus DosR/DevR box 5′-NNNGGGHCNWWNGNCCCBNN-3′ (N = any nucleotide, H = A/C/T, B = C/G/T, W = A/T) defined by Park et al. [70] and modified according to [107], [108]. Accuracy cutoff ≥85%,...
We report the genome of the facultative intracellular parasite Rhodococcus equi, the only animal pathogen within the biotechnologically important actinobacterial genus Rhodococcus. The 5.0-Mb R. equi 103S genome is significantly smaller than those of environmental rhodococci. This is due to genome expansion in nonpathogenic species, via a linear ga...
Corynebacteria grow by wall extension at the cell poles, with DivIVA being an essential protein orchestrating cell elongation and morphogenesis. DivIVA is considered a scaffolding protein able to recruit other proteins and enzymes involved in polar peptidoglycan biosynthesis. Partial depletion of DivIVA induced overexpression of cg3264, a previousl...
Rhodococcus equi is the only animal pathogen among the rhodococci. A soil inhabitant, R. equi is prevalent in the farm environment where it uses herbivore manure as growth substrate. In addition to its saprophytic lifestyle,
R. equi has the ability to colonize animal host tissues, causing pyogranulomatous infections in a variety of mammals. Althoug...
Homologues to actin are ubiquitous in nature, and actin-based cellular skeletons are crucial for the maintenance of prokaryotic and eukaryotic cellular morphology. Regarding the prokaryotes, MreB actin-homologues sustain the peptidoglycan (PG) synthesis along the lateral cell wall of most rod-shaped bacteria; FtsA actin-homologues are essential for...
Corynebacterium glutamicum is a rod-shaped actinomycete with a distinct model of peptidoglycan synthesis during cell elongation, which takes place at the cell poles and is sustained by the essential protein DivIVA(CG) (C. glutamicum DivIVA). This protein contains a short conserved N-terminal domain and two coiled-coil regions: CC1 and CC2. Domain d...
The coiled-coil protein DivIVA is a determinant of apical growth and hyphal branching in Streptomyces coelicolor. We have investigated the properties of this protein and the involvement of different domains in its essential function and subcellular targeting. In S. coelicolor cell extracts, DivIVA was present as large oligomeric complexes that were...
We identified the first enzymes that use mycothiol and mycoredoxin in a thiol/disulfide redox cascade. The enzymes are two
arsenate reductases from Corynebacterium glutamicum (Cg_ArsC1 and Cg_ArsC2), which play a key role in the defense against arsenate. In vivo knockouts showed that the genes for Cg_ArsC1 and Cg_ArsC2 and those of the enzymes of t...
The Mur ligases play an essential role in the biosynthesis of bacterial cell-wall peptidoglycan and thus represent attractive
targets for the design of novel antibacterials. These enzymes catalyze the stepwise formation of the peptide moiety of the
peptidoglycan disaccharide peptide monomer unit. MurC is responsible of the addition of the first res...
Time-lapse imaging of Streptomyces hyphae revealed foci of the essential protein DivIVA at sites where lateral branches will emerge. Overexpression experiments
showed that DivIVA foci can trigger establishment of new zones of cell wall assembly, suggesting a key role of DivIVA in directing
peptidoglycan synthesis and cell shape in Streptomyces.
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+ plasmid, typically associated with pig isolates, and compared it with that of the horse-specific vapA+ plasmid type. pVAPB1593, a circular 79,251-bp element, had the s...
Bacterial cell growth and cell division are highly complicated and diversified biological processes. In most rod-shaped bacteria, actin-like MreB homologues produce helicoidal structures along the cell that support elongation of the lateral cell wall. An exception to this rule is peptidoglycan synthesis in the rod-shaped actinomycete Corynebacteriu...
The actinomycete Corynebacterium glutamicum grows as rod-shaped cells by zonal peptidoglycan synthesis at the cell poles. In this bacterium, experimental depletion of
the polar DivIVA protein (DivIVACg) resulted in the inhibition of polar growth; consequently, these cells exhibited a coccoid morphology. This result demonstrated
that DivIVA is requi...
Of the five promoters detected for the ftsZ gene in Corynebacterium glutamicum, three were located within the coding region of the upstream ftsQ gene and two within the intergenic ftsQ-ftsZ region. The most distant ftsZ promoter showed activity in Escherichia coli and controlled high-level transcriptional expression of ftsZ in C. glutamicum. Quanti...
Analysis of the complete genome sequence of Corynebacterium glutamicum indicated that, in addition to ftsI, there are eight proteins with sequence motifs that are strongly conserved in penicillin binding proteins (PBPs): four genes that code for high-molecular-weight (HMW)-PBPs (PBP1a, PBP1b, PBP2a and PBP2b), two genes encoding low-molecular-weigh...
On May 18, 2007 this sequence version replaced gi:110162111.
The natural resistance mechanisms of corynebacteria to respond to the environments containing high levels of arsenic were successfully adopted to develop inexpensive and selective extractants for submicrogram amounts of arsenic. Kinetic and equilibrium characteristics were evaluated, and a preliminary exploration of the capability of these strains...
The actinomycete Corynebacterium amycolatum is a saprophytic bacterium usually associated with the human skin, but it is at present considered an emergent pathogen as it is isolated from nosocomial settings from samples of immunosuppressed patients. The conventional method to distinguish C. amycolatum from closely related species is mainly based on...
![Fig. 3. Resistance level to arsenite [As(III)] or arsenate [As(V)] by...](profile/Michal-Letek/publication/6735287/figure/fig3/AS:601674146136098@1520461703574/Resistance-level-to-arsenite-AsIII-or-arsenate-AsV-by-Escherichia-coli-DH5a-1_Q320.jpg)
Arsenic is an extremely toxic metalloid that, when present in high concentrations, severely threatens the biota and human health. Arsenic contamination of soil, water, and air is a global growing environmental problem due to leaching from geological formations, the burning of fossil fuels, wastes generated by the gold mining industry present in unc...
In Corynebacterium glutamicum, as in many Gram-positive bacteria, the cell division gene ftsI is located at the beginning of the dcw cluster, which comprises cell division- and cell wall-related genes. Transcriptional analysis of the cluster revealed that ftsI is transcribed as part of a polycistronic mRNA, which includes at least mraZ, mraW, ftsL,...
The genes involved in gluconate catabolism (gntP and gntK) in Corynebacterium glutamicum are scattered in the chromosome, and no regulatory genes are apparently associated with them, in contrast with the organization
of the gnt operon in Escherichia coli and Bacillus subtilis. In C. glutamicum, gntP and gntK are essential genes when gluconate is th...
Corynebacterium glutamicum is able to grow in media containing up to 12 mM arsenite and 500 mM arsenate and is one of the most arsenic-resistant microorganisms
described to date. Two operons (ars1 and ars2) involved in arsenate and arsenite resistance have been identified in the complete genome sequence of Corynebacterium glutamicum. The operons ar...
Corynebacterium glutamicum is a Gram-positive bacterium that lacks the cell division FtsA protein and actin-like MreB proteins responsible for determining cylindrical cell shape. When the cell division ftsZ gene from C. glutamicum (ftsZ(Cg)) was cloned in different multicopy plasmids, the resulting constructions could not be introduced into C. glut...
The expression of genes coding for heterologous extracellular enzymes or proteins in corynebacteria has provided new capacities
to these industrially important microorganisms, such as the use of the culture media as sources of essential amino acids and
hydrolytic enzymes that can be used as complements in animal food or for the production of enzyme...
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Question (1)
Roehampton is pleased to offer 50 VC Scholarships for PhD research areas across the university. Funding is available for UK/EU and International* students at Home/EU rates (tuition fee waiver at £4,052 and stipend at £16,057 for 2015/16) for three years full-time study (or part time equivalent for five years).
One of the projects supported by this initiative is titled "Hijacking of host cellular functions and metabolism in MRSA pathogenesis".
Our objective is to dissect the interaction of Methicillin-Resistant Staphylococcus aureus (MRSA) with mammalian host cells. We have recently discovered that the depletion of a human membrane protein called TMEM59 results in the reduced intracellular replication of MRSA, while host cell viability remains unaffected (EMBO J, 2013. 32(4):566-82). The candidate will perform a range of genomic, proteomic and metabolic profiling studies to find previously unknown factors required for the intracellular survival of MRSA. This could lead to the identification of novel host-targeted therapeutics against MRSA, which may circumvent the problem of antibiotic resistance.
The successful applicant will join the Health Sciences Research Centre (HSRC) at the University of Roehampton. The HSRC represents a multidisciplinary team with research interests on different aspects of human disease, ranging from neurological disorders, to bacterial infections or metabolic disorders. 84% of our research output is rated as internationally excellent and world leading in terms of originality, significance and rigour (REF2014).
The HSRC has well-equipped laboratories including an ultra-high performance liquid chromatography triple quadrupole mass spectrometer (UPLC-(TQ)-MS), clinical and microscopy analysers, molecular biology equipment, and cell and tissue culture facilities. Our excellent research facilities make the HSRC a stimulating environment for PhD students.
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