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Evolution of Staphylococcus aureus during human colonization and infection
Abstract
The diversification of bacterial pathogens during infection is central to their capacity to adapt to different anatomical niches, evade the host immune system, and overcome therapeutic challenges. For example, antimicrobial treatment may fail due to the development of resistance during infection, which is often accompanied by transition to a less virulent state during chronic, persistent infection. In this review, the adaptation of the major human pathogen Staphylococcus aureus to its host environment during infection will be discussed, particularly in the context of new sequencing technologies which have opened a gateway towards understanding of the molecular processes underlying those adaptations. We now have the capacity to address previously intractable questions regarding bacterial diversification during infection which will ultimately lead to enhanced understanding of pathogenesis and the nature of epidemics, and will inform the design of effective therapeutic measures.
... Compensation for ker- (20,21). If keratinocytes contribute significantly to S. aureus defense, it is likely that these organisms have acquired mechanisms to evade keratinocyte-mediated clearance (16). We postulated that MRSA USA300 evades keratinocyte-mediated clearance through the selection of toxin-deficient mutants that can persist intracellularly. ...
... Selection of agr mutants from within infected keratinocytes. S. aureus bacteria readily adapt to environmental immune pressure through the selection and proliferation of specific mutants (16). We postulated that WT organisms cleared by caspase-1dependent pyroptosis in the cytosol would be at a selective disadvantage within keratinocytes compared to the agr mutant bacteria, which lacking toxin secretion, fail to trigger inflammasomemediated clearance. ...
... Thus, finding a small population of S. aureus within human keratinocytes is not unexpected. The evolution of S. aureus during asymptomatic carriage documents ongoing adaptation of these pathogens to the host (55) and includes mutation at the agr locus (16). Clinical studies indicate that human nasal carriage of the same S. aureus strains can persist over weeks if not longer (56). ...
Unlabelled:
Skin is the most common site of Staphylococcus aureus infection. While most of these infections are self-limited, recurrent infections are common. Keratinocytes and recruited immune cells participate in skin defense against infection. We postulated that S. aureus is able to adapt to the milieu within human keratinocytes to avoid keratinocyte-mediated clearance. From a collection of S. aureus isolated from chronically infected patients with atopic dermatitis, we noted 22% had an agr mutant-like phenotype. Using several models of human skin infection, we demonstrate that toxin-deficient, agr mutants of methicillin-resistant S. aureus (MRSA) USA300 are able to persist within keratinocytes by stimulating autophagy and evading caspase-1 and inflammasome activation. MRSA infection induced keratinocyte autophagy, as evidenced by galectin-8 and LC3 accumulation. Autophagy promoted the degradation of inflammasome components and facilitated staphylococcal survival. The recovery of more than 58% agr or RNAIII mutants (P < 0.0001) of an inoculum of wild-type (WT) MRSA from within wortmannin-treated keratinocytes compared to control keratinocytes reflected the survival advantage for mutants no longer expressing agr-dependent toxins. Our results illustrate the dynamic interplay between S. aureus and keratinocytes that can result in the selection of mutants that have adapted specifically to evade keratinocyte-mediated clearance mechanisms.
Importance:
Human skin is a major site of staphylococcal infection, and keratinocytes actively participate in eradication of these pathogens. We demonstrate that methicillin-resistant Staphylococcus aureus (MRSA) is ingested by keratinocytes and activates caspase-1-mediated clearance through pyroptosis. Toxin-deficient MRSA mutants are selected within keratinocytes that fail to induce caspase-1 activity and keratinocyte-mediated clearance. These intracellular staphylococci induce autophagy that enhances their intracellular survival by diminishing inflammasome components. These findings suggest that S. aureus mutants, by exploiting autophagy, can persist within human keratinocytes.
... Staphylococcus aureus hem toplum kaynaklı enfeksiyonlara hem de hastane enfeksiyonlarına sıklıkla neden olabilen, solunum yolu, idrar yolu, ciltyumuşak doku ve kan yolu enfeksiyonları gibi çok çeşitli klinik tablolar ile seyredebilen önemli bir bakteriyel insan patojenidir (1)(2)(3)(4) . S. aureus'un çevrede yaygın olarak bulunması yanında çoğu sağlıklı bireyin deri ve mukoza zarları gibi normal florasında da bulunması potansiyel olarak ciddi enfeksiyon kaynağı olmasını kolaylaştırmaktadır (1,2,4) . ...
... Staphylococcus aureus hem toplum kaynaklı enfeksiyonlara hem de hastane enfeksiyonlarına sıklıkla neden olabilen, solunum yolu, idrar yolu, ciltyumuşak doku ve kan yolu enfeksiyonları gibi çok çeşitli klinik tablolar ile seyredebilen önemli bir bakteriyel insan patojenidir (1)(2)(3)(4) . S. aureus'un çevrede yaygın olarak bulunması yanında çoğu sağlıklı bireyin deri ve mukoza zarları gibi normal florasında da bulunması potansiyel olarak ciddi enfeksiyon kaynağı olmasını kolaylaştırmaktadır (1,2,4) . ...
... In addition, under selective pressure the bacteria will acquire genomic variation and display phenotypic changes. The acquired genomic variations may lead to (Young et al. 2012;Fitzgerald 2014;Lindsay 2014;Messina et al. 2016;Giulieri et al. 2018;Guérillot et al. 2019). Different S. aureus strains display distinct expression of virulence factors, which are the key players for survival at different body sites and pathogenesis (Zhao et al. 2019(Zhao et al. , 2020. ...
... These findings can be related to the hypothesis that over a period of time, S. aureus colonizing the human host will acquire genetic variations associated with infection at the colonization site. This will then lead to the emergence of bacteria causing infection phenotypes when an unknown trigger is perceived by the bacteria (Fitzgerald 2014). For this reason, when an infection occurs, it is interesting to know what type of host immune failures occurred to allow S. aureus invasion and to identify the nature of the unknown trigger. ...
Staphylococcus aureus is an opportunistic human pathogen, which is a leading cause of infections world-wide. The challenge in treating S. aureus infection is linked to the development of multi-drug resistant strains and the mechanisms employed by this pathogen to evade the human immune defenses. In addition, S. aureus can hide asymptomatically in particular 'protective' niches of the human body for prolonged periods of time. In the present review, we highlight recently gained insights in the role of the human gut as an endogenous S. aureus reservoir next to the nasopharynx and oral cavity. In addition, we address the contribution of these ecological niches to staphylococcal transmission, including the roles of particular triggers as modulators of the bacterial dissemination. In this context, we present recent advances concerning the interactions between S. aureus and immune cells to understand their possible roles as vehicles of dissemination from the gut to other body sites. Lastly, we discuss the factors that contribute to the switch from colonization to infection. Altogether, we conclude that an important key to uncovering the pathogenesis of S. aureus infection lies hidden in the endogenous staphylococcal reservoirs, the trafficking of this bacterium through the human body, and the subsequent immune responses.
... S taphylococcus aureus is a commensal colonizer of 20 to 30% of healthy people as normal flora of the nasopharynx, skin, and other secondary niches (1). Colonization leads to elevated risk for metastatic S. aureus infections of the host. ...
... Colonization leads to elevated risk for metastatic S. aureus infections of the host. S. aureus strains cause a large diversity of infections (1,2). Factors underpinning this diversity include robust stress resilience, biofilm formation, resistance to successive antibiotics, generation of antibiotic-tolerant persisters, and evasion of the host immune response (3)(4)(5)(6). ...
Staphylococcus aureus has three types of cation/proton antiporters. The type 3 family includes two multisubunit Na⁺/H⁺ (Mnh) antiporters, Mnh1 and Mnh2. These antiporters are clusters of seven hydrophobic membrane-bound protein subunits. Mnh antiporters play important roles in maintaining cytoplasmic pH in prokaryotes, enabling their survival under extreme environmental stress. In this study, we investigated the physiological roles and catalytic properties of Mnh1 and Mnh2 in S. aureus. Both Mnh1 and Mnh2 were cloned separately into a pGEM3Z+ vector in the antiporter-deficient KNabc Escherichia coli strain. The catalytic properties of the antiporters were measured in everted (inside out) vesicles. The Mnh1 antiporter exhibited a significant exchange of Na⁺/H⁺ cations at pH 7.5. Mnh2 showed a significant exchange of both Na⁺/H⁺ and K⁺/H⁺ cations, especially at pH 8.5. Under elevated salt conditions, deletion of the mnhA1 gene resulted in a significant reduction in the growth rate of S. aureus in the range of pH 7.5 to 9. Deletion of mnhA2 had similar effects but mainly in the range of pH 8.5 to 9.5. Double deletion of mnhA1 and mnhA2 led to a severe reduction in the S. aureus growth rate mainly at pH values above 8.5. The effects of functional losses of both antiporters in S. aureus were also assessed via their support of virulence in a mouse in vivo infection model. Deletion of the mnhA1 gene led to a major loss of S. aureus virulence in mice, while deletion of mnh2 led to no change in virulence.
... Asimismo, está bien entendido que Staphylococcus aureus tiene una habilidad notoria para volverse resistente a los antibióticos en poco tiempo ya que posee una alta tasa de mutación y además adquiere fácilmente elementos genéticos determinantes de resistencia (Chambers & Deleo, 2009;Fitzgerald, 2014;Kobayashi & DeLeo, 2009). Esto ocasiona que su manejo clínico sea complejo [Ver Figura 2]. ...
... Finalmente, es importante comprender que S. aureus tiene la propiedad biológica fundamental de ser flora normal del organismo: aproximadamente 30% de los humanos son transportadores nasales no sintomáticos de S. aureus (Kluytmans, van Belkum, & Verbrugh, 1997). Específicamente, se puede encontrar S. aureus en la nasofaringe, la piel, la garganta, y el tracto gastrointestinal de las personas sanas (asintomáticas), así como en el tracto urogenital de las mujeres (Fitzgerald, 2014). Esto ocasiona que el microorganismo esté expuesto a un sinnúmero de nuevos antibióticos que la población se automedica indiscriminadamente, especialmente en Colombia. ...
Antibacterial resistance is a serious problem of public health in Colombia and the world. There are various approaches to mitigate this growing problem. Among them, is to increase the antibiotic effectiveness of drugs through a proper delivering in pharmaceutical matrices; for example, the association of antimicrobials in polymeric pseudophases that act as bioprotector reservoirs against enzymatic degradation. This project had as general objective the evaluation of antimicrobial activity of beta-lactam antibiotic ampicillin associated with the amphiphilic polymer matrix PAM-18 disodium in three strains of Staphylococcus aureus, with different degrees of susceptibility to this type of antibiotics.
Initially, the use of UV-Vis spectrophotometry was evaluated for the determination of the minimum inhibitory concentration in broth microdilution tests. For this, a bacterial quantification protocol was done, in which it was obtained calibration curves that relate the absorbance measured with the counts of CFU/mL. However, these curves were inadequate by the low presented linearity and narrow absorbance evaluated interval; finally, they were not used in the analysis of the data obtained in subsequent tests and require its repetition with an improved design of the trial. In addition, it was concluded that, due to the visual patterns of growth that showed the strains under study, it was inaccurate to quantify bacterial growth using UV-Vis spectrophotometry. In contrast, the evaluated inspection methods (visual and spectrophotometric) had a proper performance for qualitatively estimating bacterial growth and it was presented both its advantages and disadvantages according to the objective of the project.
Subsequently, it was determined the in vitro minimum inhibitory concentrations of various treatments (polymer, ampicillin and complex association of different proportions of the components) in each of the strains that were under study, by conducting thirteen antimicrobial susceptibility tests using two robust and reliable methodologies of broth microdilution based on the criteria of CLSI. It was confirmed that the polymer PAM-18 disodium doesn’t exert antibiotic effect against the strains that were studied. The need for the vortex to generate the correct formation of the complex association was highlighted. The strains under study presented variables MIC toward ampicillin. The most effective treatment to inhibit the growth of S. aureus ATCC ® 25923 corresponded to the association complex A:P 1:1 V, because the MIC of this treatment was slightly lower compared to free ampicillin treatment (change of 0.125 μg/mL to 0.06 μg/mL). For resistant strains S. aureus ATCC ® 29213 and S. aureus ATCC® 43300, it was not clearly revealed a bioprotective effect of the complex association compared to the free drug, since MIC were similar for all treatments.
Although previous studies showed that the polymer formed hydrophobic pseudophases capable of retaining ampicillin due to intermolecular forces, several factors could affect the expected bioprotective performance of the matrix: the association complexes were not formed or were unstable under the conditions studied; the drug was adsorbed at the interface, and not in the core, of the coiled polymer and thus exposed to enzymatic degradation; there were unfavorable electrostatic or steric interactions between the polymer and the cell wall; ampicillin was not completely protected until its arrival at the target or the polymer suffered other kinds of relations with the bacteria such as endocytosis. Thus, future research that elicits the physicochemical and molecular aspects of the evaluated phenomenon is required.
... These new broad-range genetic techniques may provide a basis for future virulence assignment of clinical isolates (Laabei et al., 2014). MRSA virulence can depend on known and presumably also on still unknown virulence determinants apart from environmental and regulatory processes (Fitzgerald, 2014). Genotypic differences between sole commensal and invasivecompetent MRSA strains can be investigated in particular by focusing isolates of the same CC (Gordon and Lowy, 2008). ...
... WGS of the RN4220 laboratory strain pointed out that there are a number of SNPs affecting the general fitness of S. aureus (Nair et al., 2011). Most relevant to the present work were the recent WGS studies which were able to associate the toxicity of 90 MRSA ST239 isolates with around 100 statistically significant SNPs (Laabei et al., 2014) and studies investigating the evolutionary dynamics of S. aureus during long-term carriage and transition from nasal carriage to invasive infection (Chua et al., 2014;Fitzgerald, 2014;Golubchik et al., 2013;Young et al., 2012). ...
Hospital-associated methicillin-resistant Staphylococcus aureus (MRSA) infections are frequently caused by predominant clusters of closely related isolates that cannot be discriminated by conventional diagnostic typing methods. Whole genome sequencing (WGS) and DNA microarray (MA) now allow for better discrimination within a prevalent clonal complex (CC). This single center exploratory study aims to distinguish invasive (blood stream infection) and non-invasive (nasal colonization) MRSA isolates of the same CC5 into phylogenetic- and virulence-associated genotypic subgroups by WGS and MA. A cohort of twelve blood stream and fifteen nasal MRSA isolates of CC5 (spa-types t003 and t504) was selected. Isolates were propagated at the same period of time from unrelated patients treated at the University of Saarland Medical Center, Germany. Rooted phylotyping based on WGS with core-genome single nucleotide polymorphism (SNP) analysis revealed two local clusters of closely related CC5 subgroups (t504 and Clade1 t003) which were separated from other local t003 isolates and from unrelated CC5 MRSA reference isolates of German origin. Phylogenetic subtyping was not associated with invasiveness when comparing blood stream and nasal isolates. Clustering based on MA profiles was not concordant with WGS phylotyping, but MA profiles may identify subgroups of isolate with nasal and blood stream origin. Among the new putative virulence associated genes identified by WGS, the strongest association with blood stream infections was shown for ebhB mutants. Analysis of the core-genome together with the accessory genome enables subtyping of closely related MRSA isolates according to phylogeny and presumably also to the potential virulence capacity of isolates.
Copyright © 2015. Published by Elsevier B.V.
... Staphylococcus aureus (S. aureus) is a commensal opportunistic gram-positive bacterium that colonizes around 20-30% of people, but it can become pathogenic for the presence of virulence and immune evasion factors, causing different diseases ranging from mild skin infections to toxic shock syndrome (TSS) and multi-organ failure (1)(2)(3)(4). The bacterium is a component of the nasopharynx microbiota that may colonize secondary sites including skin, throat, armpit, groin and intestine (2), where it can survive thanks to factors and enzymes that allow its adherence to the epithelial surfaces and proliferation (5). The persistent proliferation and survival of S. aureus at the site of infection often lead to dysbiosis with serious life-threatening conditions, including chronic inflammation (6). ...
Staphylococcus aureus is a gram-positive bacterium that may cause intestinal inflammation by secreting enterotoxins, which commonly cause food-poisoning and gastrointestinal injuries. Staphylococcal enterotoxin B (SEB) acts as a superantigen (SAg) by binding in a bivalent manner the T-cell receptor (TCR) and the costimulatory receptor CD28, thus stimulating T cells to produce large amounts of inflammatory cytokines, which may affect intestinal epithelial barrier integrity and functions. However, the role of T cell-mediated SEB inflammatory activity remains unknown. Here we show that inflammatory cytokines produced by T cells following SEB stimulation induce dysfunctions in Caco-2 intestinal epithelial cells by promoting actin cytoskeleton remodelling and epithelial cell-cell junction down-regulation. We also found that SEB-activated inflammatory T cells promote the up-regulation of epithelial-mesenchymal transition transcription factors (EMT-TFs) in a nuclear factor-κB (NF-κB)- and STAT3-dependent manner. Finally, by using a structure-based design approach, we identified a SEB mimetic peptide (pSEB116-132) that, by blocking the binding of SEB to CD28, dampens inflammatory-mediated dysregulation of intestinal epithelial barrier.
... Colonization includes superficial residence of these bacteria on the skin as a component of the microbiome (Oh et al., 2016), intercalation within the layers of the epidermis (Nakatsuji et al., 2013), and intracellular persistence within keratinocytes (Soong et al., 2015). Skin and soft-tissue infections, as well as systemic S. aureus infection, are usually caused by colonizing strains, well documented in multiple whole-genome studies (Fitzgerald, 2014). Although typically considered to be extracellular pathogens, S. aureus can also persist within immune, endothelial, and epithelial cells that can serve as foci of persistent or recurrent infection. ...
OBJECTIVE
Human skin is commonly colonized and infected by Staphylococcus aureus . Under normal conditions skin is relatively hypoxic. Thus, we postulated that the burden of staphylococcal infection would result in competition for glucose and stimulation of HIF1a signaling to initiate keratinocyte metabolic and immune responses to infection.
METHODS
The metabolic activity of primary keratinocytes in response S. aureus metabolic mutants was monitored using a SeaHorse analyzer to measure oxygen consumption and extracellular acidification. A transcriptomic analysis of keratinocyte responses to S . aureus USA300 infected keratinocytes was performed and key results were confirmed by qRT‐PCR and immunoblot. The immunological consequences of Hif1a signaling were demonstrated in vitro, as the metabolic and inflammatory activities of primary keratinocytes were studied through qRT arrays, western blots, ELISA assays and confocal imaging. The in vivo consequences of the glycolytic response to infection were confirmed in vivo in a murine model of skin infection.
ABSTRACT
Using a combination of metabolic and transcriptomic methodologies, we found that S. aureus infection is sensed as a metabolic stress by keratinocytes. This is sufficient to induce HIF1a signaling, which promotes IL‐1b production and generates components of the aerobic glycolysis pathway to meet the metabolic requirements of infection. Staphylococci capable of glycolysis, including WT and agr mutants readily induced HIF1a. In contrast, pyk glycolytic mutants failed to compete with keratinocytes for their metabolic needs. Although S. aureus activate the Akt/mTOR cascade, inhibition of this pathway did not inhibit the keratinocyte cytokine response to infection. However, 2‐DG mediated suppression of glycolysis blocked keratinocyte production of IL‐1b in vitro and significantly exacerbated the S. aureus cutaneous infection in a murine model.
CONCLUSIONS
S. aureus is perceived as a metabolic stress by infected keratinocytes. These keratinocytes respond through HIF1a signaling, increasing glycolytic activity and generating IL‐1b. S. aureus glycolytic mutants, independent of toxin production, fail to activate keratinocytes.
Support or Funding Information
FUNDING: The results presented in this abstract were supported by the National Institutes of Health under award number RO1AI103854.
... In humans, the most relevant species is S. aureus because is responsible for many nosocomial infections that can risk a patient's life, especially in the case of antibiotic-resistant strains. This bacterium can also harm animals, together with the Staphylococcus hyicus and Staphylococcus intermedius group, even though these have a limited host range compared to S. aureus [84,85]. None of the sampled animals were positive for S. aureus, which matches with other studies in reptiles such as Espinosa-Gongora et al. [86], in which no bacteria were isolated in any of the 21 samples examined from multiple reptile species, or Cristina et al. [70] work, with a S. aureus prevalence of 2.5% (1/43). ...
Veiled chameleons are native to the Arabian Peninsula, with a presence in other regions, such as the Canary Islands (Spain). The aim of this study is to analyze the existence of pathogenic bacteria in a population of this invasive reptile on Gran Canaria island. The results obtained highlight the presence of a variety of pathogens with relevance to human health, most of them related to gastrointestinal diseases. This archipelago is a biodiversity hotspot, with some endangered species living there, so the presence of veiled chameleons could be also a risk to biodiversity
conservation, by the spread and/or transmission of pathogenic bacteria to the native fauna. In conclusion, the invasive veiled chameleon population in the Canary Islands should be considered as a potential risk factor for biodiversity conservation and human health.
... To date, the literature lacks information regarding bacterial colonization on titanium microplates used in oral and maxillofacial surgery. However, multiple studies have investigated the microbiological and biofilm characteristics of bacteria on medical devices implanted in the body or on human tissue [20]. ...
The present investigation was undertaken to evaluate the topographical and ultrastructural architecture of titanium plates coated with polylactic co-glycolic acid (PLGA), chitosan (CH), and/or meropenem (MEM) with or without Staphylococcus aureus (SA) or Pseudomonas aeruginosa (PA) bacteria. Single-hole segments of 0.4 mm thick, low-profile titanium plates were spray coated using an airbrush with polymeric carriers (PLGA or CH) loaded with MEM, in addition to the negative control group (uncoated titanium plates). The coated plates and the negative control group were subjected to bacterial biofilms through a cultivation process while being slowly stirred at 20 rpm for 24 h. The samples were fixed and processed for scanning electron microscopic study at 5, 10, and 20 k magnification. The data were statistically analyzed to compare within and between the different materials. Coating titanium plates with PLGA or CH with MEM appeared to enhance bacterial inhibition over uncoated plates, hindering biofilm formation and preventing bacterial proliferation. In the staphylococcus aureus group, the highest bacterial count was observed in the uncoated plates, whereas the lowest count was detected in meropenem-PLGA, followed by PLGA, chitosan, meropenem, and meropenem-chitosan, respectively. On the other hand, the Pseudomonas aeruginosa group with the uncoated plates had the highest bacterial count, whereas the lowest bacterial count was found related to CH, followed by PLGA, MP, MC, and MEM, respectively.
... Many strains of S. aureus have become resistant to antibiotics notably methicillin and they are regarded as methicillin-resistant S. aureus (MRSA) (Fitzgerald, 2014). The resistance to methicillin and other β-lactam antibiotics is due to the possession of mecA gene, which is found on a mobile genetic element, the staphylococcal cassette chromosome (SCCmec). ...
The resistance of bacteria to some antibiotics is rampant nowadays. One of the genera of bacteria that has shown resistance to notable antibiotics is Staphylococcus aureus. Therefore, this study aimed at isolation of S. aureus from the soils and wooden table surfaces in some abattoirs within Ilorin, Nigeria. The objectives of this study were to determine the counts of heterotrophic bacteria in the soils and wooden table surfaces at some abattoirs in Ilorin, Nigeria; characterize and identify the S. aureus isolates; determine the antibiotic resistance patterns of the isolates; and detect the presence of spa, mecA, and mecC in the isolates. The counts of viable bacteria and S. aureus, antibiotic susceptibility as well as the physicochemical characteristics were determined using standard methods. The heterotrophic bacterial counts of the soils and wooden table surfaces ranged from 1.0 x 105 – 3.4 x 107 and 2.1 x 105 – 1.0 x 108 cfu/g respectively. In addition, the counts of S. aureus in the soils and wooden table surfaces ranged from 1.0 x 103 – 3.0 x 107 and 1.0 x 103 – 3.1 x 106 cfu/g respectively. The S. aureus isolates were positive for Gram staining, catalase, oxidase, mannitol fermentation and coagulase test. All the S. aureus were resistant to ampicillin, and ceftazidime, followed by augmentin (82%), vancomycin, cefuroxime, and nitrofurantoin (55% each); gentamicin and ciprofloxacin (36%) and ofloxacin (18%). None of the isolates possess mecA gene while mecC and spa genes were detected in 45.5 and 54.5% of the isolates respectively. This is the first study in which mecC gene was detected in any S. aureus in Nigeria. It is concluded from this study that the soils and wooden table surfaces from the abattoirs harboured S. aureus some of which possessed mecC for antibiotic resistance and the virulent factor, spa genes.
... Of the 25 Staphylococcus isolates collected in the present study, all but the four S. pseudintermedius strains were of a typically human origin [12][13][14], even though the S. aureus strains were isolated from both humans and the family dog. On the contrary, S. pseudintermedius, typically colonizing dogs' skin [13], was isolated during the entire sampling time span from the dog only. ...
Although usually harmless, Staphylococcus spp. can cause nosocomial and community-onset skin and soft tissue infections in both humans and animals; thus, it is considered a significant burden for healthcare systems worldwide. Companion animals have been identified as potential reservoirs of pathogenic Staphylococcus with specific reference to Methicillin Resistant Staphylococcus aureus (MRSA). In this study, we investigated the circulation and the genetic relationships of a collection of Staphylococcus spp. isolates in a family composed of four adults (a mother, father, grandmother, and grandfather), one child, and a dog, which were sampled over three years. The routes of transmission among humans and between humans and the dog werelyzed. The results displayed the circulation of many Staphylococcus lineages, belonging to different species and sequence types (ST) and being related to both human and pet origins. However, among the observed host-switch events, one of them clearly underpinnthroponotic route from a human to a dog. This suggests that companion animals can potentially have a role as a carrier of Staphylococcus, thus posing a serious concern about MRSA spreading within human and animal microbial communities.
... S taphylococcus aureus is a gram-positive opportunistic pathogen most frequently isolated from community-and hospital-acquired infections, and is one of the most significant clinical pathogens, because of its capacity to adapt to diverse environmental conditions. [1][2][3] Methicillin-resistant Staphylococcus aureus (MRSA) refer to those S. aureus strains that express the mecA gene or other resistance mechanisms, and have also shown cefoxitin (FOX) or oxacillin (OXA) resistance 4,5 by antibiotic susceptibility testing (AST). Infections caused by MRSA are more difficult to treat than that of methicillin-susceptible S. aureus. ...
Methicillin-resistant Staphylococcus aureus (MRSA) are challenging pathogenic bacteria that can cause severe infection leading to high mortality rates. We found that both the oxacillin- and cefoxitin-resistant S. aureus strains isolated from clinic showed multidrug-resistant (MDR) characteristics. Through rapid high-throughput screen (HTS) of a compound library, gemcitabine and selen compounds were found to effectively inhibit S. aureus growth. For further improvement, we synthesized selen-containing gemcitabine that demonstrated relatively potent antimicrobial activity against several MDR MRSA in vitro. The HTS assay and gemcitabine selen derivative provided a useful tool to explore an effective molecular target to treat MDR MRSA.
... The innate adaptability of S. aureus has led to the emergence of resistance to multiple classes of antibiotics through the acquisition of mobile genetic elements (MGE) encoding resistance determinants, or mutations in loci influencing antibiotic sensitivity Jensen and Lyon 2009;Fitzgerald 2014). Methicillin-resistant S. aureus (MRSA) have been documented at a rapid and increasing rate since methicillin was first introduced in 1959. ...
Staphylococcus aureus is an opportunistic pathogen and one of the major bacteria responsible for community-acquired and nosocomial infections for which the treatment is complicated by the emergence of multidrug resistant strains. Its adaptation to multiple growth conditions, which contributes to its virulence, is under the control of numerous factors including regulatory RNAs, often called sRNAs for small RNAs. We performed an accurate survey of all sRNAs from the model strain HG003, a NCTC8325 derivative commonly used for S. aureus genetic regulation studies. It is a complex task, essential to accomplish molecular and functional studies. We found about 50 bona fide sRNAs, a number much lower than previously reported.As most sRNAs contribute to the "fine-tuning" of gene expression, sRNA-dependent phenotypes are generally difficult to detect. However, sRNA-mediated phenotypes may emerge as dominant traits after a few generations under selective pressure. We set up an experimental strategy to evaluate the fitness of S. aureus sRNA mutants within a population of sRNA mutants. A library of eighty HG003 mutants with sRNA genes replaced by specific DNA “barcode” sequences to allow the identification of each mutant was constructed. The mutant library was grown in different conditions, DNA barcodes were PCR and counted by DNA-seq. Mutants diminishing or accumulating during a stress condition provide information on sRNA functions. The use of specific primers allows multiplexing 50 experimental conditions in one DNA-seq library. We questioned whether sRNAs could affect the antibiotic resistance in S. aureus. Here, we present data using the above-described method with the sRNA mutant library tested in the presence of 10 antibiotics (targeting the cell wall and cell membrane, inhibiting the biosynthesis of protein, interrupting DNA replication and RNA synthesis of bacteria). Several mutants were affected by the tested growth conditions. For examples, the proportion of mutant sau6836 increases dramatically in vancomycin and reduces in flucloxacillin, cloxacillin and cefazolin. The proportion of RNAIII-agr mutant increases progressively in the presence of gentamicin, linezolid and clindamycin. The proportion of 6S RNA mutant decreases significantly in the presence of rifampicin. Interestingly, the 6S RNA and the rifampicin are targeting the RNA polymerase. The sRNA RsaA targets a regulator of autolytic activities and its corresponding mutant is remarkably reduced in the presence of ciprofloxacin. These examples illustrate the power of fitness experiments to identify phenotypes and reveals that several sRNAs contribute to modulate the resistance to antibiotics.
... They reflect an adaptive strategy developed by CF pathogens for long-term survival in the CF lung, allowing them to quickly adapt and face fluctuating selective pressures, the so-called "insurance hypothesis" [35]. For S. aureus, diversified populations have been more rarely documented in CF [14,[36][37][38][39] and population heterogeneity has only once been related to antimicrobial susceptibility patterns [36]. Here, we documented intrasample heterogeneity of susceptibility to linezolid, which may explain some of the previous observations (alternate detection of susceptible and resistant strains, non-chronological accumulation of SNPs) made by other authors [14,17,23]. ...
Methicillin-resistant Staphylococcus aureus (MRSA) can cause chronic lung infections in patients with Cystic Fibrosis (CF). One option for managing them is the use of linezolid. We hereby report the in-host emergence of linezolid resistance (LR) in MRSA in CF siblings via a population analysis. A collection of 171 MRSA strains from 68 samples were characterized by determining their linezolid Minimal Inhibitory Concentrations (MICs), analyzing the locus of staphylococcal protein A (spa) and whole genome sequencing. Courses of linezolid were retraced. Strains belonged to three spa types (t002, t045, t127) and two sequence types (ST1, ST5). Emergence of LR occurred under treatment, one year apart in both siblings, in the CC5-MRSA-I Geraldine clone harboring the toxic shock syndrome toxin-1-encoding gene. Resistance was related to a G2576T substitution present in a variable number of 23S rRNA gene copies. Susceptible and resistant strains were co-isolated within samples. Single Nucleotide Polymorphism-based analysis revealed complex colonizations by highly diversified, clonally related populations. LR remains rare in MRSA and there are very few longitudinal analyses documenting its emergence. Analyzing a large MRSA collection revealed new aspects of LR emergence: it emerges in specific subclonal lineages resulting from adaptive diversification of MRSA in the CF lung and this heterogeneity of intra-sample resistance may contribute to compromising antibiotic management.
... Humans are a natural reservoir of S. aureus. Between 30% and 50% of healthy adults are colonized, and between 10% and 20% permanently colonized [97,98]. S. aureus is a commensal organism and also an important opportunistic human pathogen, causing a variety of community and hospital-associated pathologies, including bacteremia-sepsis, endocarditis, pneumonia, osteomyelitis, arthritis, and skin diseases [97,99]. ...
Infectious diseases occur worldwide with great frequency in both adults and children. Both infections and their treatments trigger mitochondrial interactions at multiple levels: (i) incorporation of damaged or mutated proteins to the complexes of the electron transport chain, (ii) mitochondrial genome (depletion, deletions, and point mutations) and mitochondrial dynamics (fusion and fission), (iii) membrane potential, (iv) apoptotic regulation, (v) generation of reactive oxygen species, among others. Such alterations may result in serious adverse clinical events with great impact on children’s quality of life, even resulting in death. As such, bacterial agents are frequently associated with loss of mitochondrial membrane potential and cytochrome c release, ultimately leading to mitochondrial apoptosis by activation of caspases-3 and -9. Using Rayyan QCRI software for systematic reviews, we explore the association between mitochondrial alterations and pediatric infections including (i) bacterial: M. tuberculosis, E. cloacae, P. mirabilis, E. coli, S. enterica, S. aureus, S. pneumoniae, N. meningitidis and (ii) parasitic: P. falciparum. We analyze how these pediatric infections and their treatments may lead to mitochondrial deterioration in this especially vulnerable population, with the intention of improving both the understanding of these diseases and their management in clinical practice
... This bacterium also has the potential to form biofilms, making it even harder to treat. S. aureus is a Grampositive coccoid commensal bacterium of the microbiota of the human body; however, some strains of S. aureus are associated with severe infections in humans [190]. Bloodstream infections in HAI are associated with catheter usage in 37% of all cases. ...
Nosocomial infections (NIs) pose an increasing threat to public health. The majority of NIs are bacterial, fungal, and viral infections; however, parasites also play a considerable role in NIs, particularly in our increasingly complex healthcare environment with a growing proportion of immunocompromised patients. Moreover, parasitic infections acquired via blood transfusion or organ transplantation are more likely to have severe or fatal disease outcomes compared with the normal route of infection. Many of these infections are preventable and most are treatable, but as the awareness for parasitic NIs is low, diagnosis and treatment are often delayed, resulting not only in higher health care costs but, importantly, also in prolonged courses of disease for the patients. For this article, we searched online databases and printed literature to give an overview of the causative agents of parasitic NIs, including the possible routes of infection and the diseases caused. Our review covers a broad spectrum of cases, ranging from widely known parasitic Nis, like blood transfusion malaria or water-borne cryptosporidiosis, to less well-known Nis, such as the transmission of Strongyloides stercoralis by solid organ transplantation or nosocomial myiasis. In addition, emerging NIs, such as babesiosis by blood transfusion or person-to-person transmitted scabies, are described.
... Apart from S. aureus, many coaqulase-negative staphylococci species are also disease-inducible in humans, of which S. epidermis is the most common pathogen of primary bacteria and infectious of indwelling medical devices [8,9]. Having a strong antimicrobial resistance, S. epidermis causes significant problems in many clinical settings and the antibiotic-resistant strains are classified as a 'High Priority' pathogen [10,11]. In general, staphylococci are able to perform genome evolution in which they alter their genome constantly to adapt to new environments, giving rise to the emergence of increasingly successfully, antibiotic-resistant, immune-evading, and host-adapted pathogens [12]. ...
Given the high therapeutic value of the staphylococcal phage, the genome co-evolution of the phage and the host has gained great attention. Though the genome-wide AT richness in staphylococcal phages has been well-studied with nucleotide usage bias, here we proved that host factor, lifestyle and taxonomy are also important factors in understanding the phage nucleotide usages bias using information entropy formula. Such correlation is especially prominent when it comes to the synonymous codon usages of staphylococcal phages, despite the overall scattered codon usage pattern represented by principal component analysis. This strong relationship is explained by nucleotide skew which testified that the usage biases of nucleotide at different codon positions are acting on synonymous codons. Therefore, our study reveals a hidden relationship of genome evolution with host limitation and phagic phenotype, providing new insight into phage genome evolution at genetic level.
... Ce sont les EGM qui participent le plus à la diversité génétique de l'espèce S. aureus, conférant un avantage sélectif aux souches qui les possèdent, comme la résistance aux antibiotiques, la résistance aux stress environnementaux ou la capacité de coloniser de nouveaux hôtes (Bosi et al., 2016). De nombreux facteurs de virulence sont sur des EGM et leur transfert horizontal au sein de l'espèce entraîne une augmentation de la pathogénicité des souches de S. aureus (Fitzgerald, 2014). ...
Staphylococcus aureus (S. aureus) est un pathogène humain majeur dont l’impact sur la santé publique est majoré par les phénomènes de résistance et de tolérance aux antibiotiques. Au cours de cette thèse, nous avons identifié deux nouveaux systèmes toxine-antitoxine (STA) de type I appartenant au core génome et exprimés par S. aureus nommés sprG2/SprF2, sprG3/SprF3. Ces nouveaux STA sont homologues au STA sprG1/SprF1 localisé dans un îlot de pathogénie. Nous avons montré que des interactions croisées influençant le niveau des ARN sprG et SprF pouvaient avoir lieu entre les STA homologues sprG/SprF, mais que celles-ci n’avaient pas d’impact sur la neutralisation spécifique de chaque toxine SprG par son antitoxine SprF. Nous avons démontré que les peptides encodés par sprG2 et sprG3 sont bactériostatiques, contrairement aux peptides encodés par sprG1 qui sont bactéricides. Nous avons démontré que l’expression des ARN sprG et SprF pouvait varier en réponse à des stress environnementaux comme un stress hyperosmotique ou un stress oxydatif. Pour le STA sprG1/SprF1, nous avons démontré que l’antitoxine SprF1 pourrait participer à l’entrée en persistance de S. aureus, en atténuant la traduction globale via son association aux ribosomes. Ainsi, SprF1 est le premier exemple d’une antitoxine ARN avec une double fonction de neutralisation de la toxine sprG1 via son extrémité 3’ et de fixation aux ribosomes pour atténuer la traduction de S. aureus via son extrémité 5’. Ensemble, ces travaux de thèse suggèrent que les STA sprG/SprF seraient impliqués dans l’adaptation de S. aureus au stress antibiotique ou dans l’échappement au système immunitaire.
... D'autres exemples d'études de l'évolution adaptative bactérienne au travers des colonisations chroniques ont porté sur la bactérie Staphylococcus aureus, qui a la capacité de coloniser de multiples niches chez l'homme. Selon la niche occupée, cette bactérie pourra être portée de manière asymptomatique, par exemple dans la zone naso-pharyngée, ou provoquer des infections chroniques persistantes, par exemple dans les voies respiratoires des patients atteints de mucoviscidose 94 . D'autres pathogènes sont par ailleurs capables de provoquer des infections pulmonaires chroniques chez les patients mucoviscidosiques, qui deviennent malgré eux un modèle pour l'étude de l'évolution adaptative bactérienne in situ. ...
L’infection pulmonaire chronique à Pseudomonas aeruginosa (P. a.) est considérée comme la principalecause de morbidité et de mortalité liée à la mucoviscidose. Au cours de cette infection persistante, labactérie s'adapte à l’environnement pulmonaire caractéristique de ces patients et évolue avec son hôtependant des décennies. Cette évolution adaptative est portée par les phénotypes, avec notamment unediminution de la virulence et une augmentation de la résistance aux antibiotiques au cours du temps. Bienque plusieurs études aient tenté d’évaluer les mécanismes génétiques de cette évolution, il demeureaujourd’hui difficile d’expliquer les relations entre les mutations accumulées dans le génome bactérien etl’expression de phénotypes cliniquement pertinents, ou encore de corréler ces mutations avec l’état desanté du patient.Nous proposons dans ce travail d’étudier les mécanismes sous-tendant cette évolution adaptative à unniveau d’observation post-génomique : la métabolomique. Dernière-née des disciplines –omiques, lamétabolomique permet la prise de vue instantanée du métabolisme, et offre une vision au plus proche duphénotype. Pour cela, nous avons constitué une banque de lignées clonales évolutives de P. a. prélevéesau cours de l’infection pulmonaire chronique chez des patients atteints de mucoviscidose. Cette banque aensuite été caractérisée aux plans clinique, phénotypique et métabolomique. L’intégration de ces différentsniveaux d’information par des méthodes statistiques multi-tableaux nous a permis de mettre en évidencedes voies métaboliques impliquées dans la patho-adaptation de P. a. à son hôte.Nos résultats permettent de faire émerger de nouvelles hypothèses pour le développement d’outilsthérapeutiques et diagnostiques visant à améliorer la prise en charge de ces infections particulièrementrésistantes aux antibiotiques. De plus, nos travaux démontrent l’intérêt de la métabolomique pour l’étudede l’évolution adaptative bactérienne en conditions naturelles.
... Understanding the genetic mechanisms of host adaptation is essential to identify novel molecular targets for tackling cross-species infections and preventing emerging clones. However, our knowledge of the population dynamics and host-adaptive genetic events that underpin these host-switching events is limited (6). To date, several studies have compared the genomes of contemporary clinical isolates from different host species and revealed genetic signatures associated with host adaptation (2,5,7). ...
While many bacterial pathogens are restricted to single host species, some have the capacity to undergo host switches, leading to the emergence of new clones that are a threat to human and animal health. However, the bacterial traits that underpin a multihost ecology are not well understood. Following transmission to a new host, bacterial populations are influenced by powerful forces such as genetic drift that reduce the fixation rate of beneficial mutations, limiting the capacity for host adaptation. Here, we implement a novel experimental model of bacterial host switching to investigate the ability of the multihost pathogen Staphylococcus aureus to adapt to new species under continuous population bottlenecks. We demonstrate that beneficial mutations accumulated during infection can overcome genetic drift and sweep through the population, leading to host adaptation. Our findings highlight the remarkable capacity of some bacteria to adapt to distinct host niches in the face of powerful antagonistic population forces.
... Because of which they are passed throught human population. Therefore fitness of them as species goes on increasing [4] S. aureus shows skin infections, food poisoning, cellulitis, bone and joint infections, bacteremia (bloodsteam infection), animal infections in cats, horses, dogs, chickens, and cows. They were found in the form of biofilm [5]. ...
Sonicated sol-gel method was used to prepare LaNiO3 from lanthanum nitrate hexahydrate La(NO3)3.6H2O(LN), nickel nitrate Ni(NO3)3.6H2O(NN), glycine and urea. Nanocrystalline LaNiO3 powder was formed after heating at 175 °C in 5 min. Particle size of LaNiO3 nanopowder was determined by Debay Scherrer’s equation and was found 48 nm. Prepared nanocatalyst characterized with the help of XRD, TGA, SEM, IR, BET surface area, EDX. Surface area of LaNiO3 was 9.22 m²/g. We have reported first time good antibacterial activity of LaNiO3 for Staphylococcus aureus. Zone of inhibition for LaNiO3 was 13 mm studied with the help of agar cup method.
... It was proposed that a diminished SigB-functionality might not be detrimental 28 , and that the complex, fine-tuned regulatory SigB network rather is modulating virulence than determining it 47 . However, we found a clear trend towards reduced virulence of the HA isolate, as one would expect in chronic, persistent infections 48 . ...
Staphylococcus aureus is a major cause of bovine mastitis, commonly leading to long-lasting, persistent and recurrent infections. Thereby, S. aureus constantly refines and permanently adapts to the bovine udder environment. In this work, we followed S. aureus within-host adaptation over the course of three months in a naturally infected dairy cattle with chronic, subclinical mastitis. Whole genome sequence analysis revealed a complete replacement of the initial predominant variant by another isogenic variant. We report for the first time within-host evolution towards a sigma factor SigB-deficient pathotype in S. aureus bovine mastitis, associated with a single nucleotide polymorphism in rsbU (G368A → G122D), a contributor to SigB-functionality. The emerged SigB-deficient pathotype exhibits a substantial shift to new phenotypic traits comprising strong proteolytic activity and poly-N-acetylglucosamine (PNAG)-based biofilm production. This possibly unlocks new nutritional resources and promotes immune evasion, presumably facilitating extracellular persistence within the host. Moreover, we observed an adaptation towards attenuated virulence using a mouse infection model. This study extends the role of sigma factor SigB in S. aureus pathogenesis, so far described to be required for intracellular persistence during chronic infections. Our findings suggest that S. aureus SigB-deficiency is an alternative mechanism for persistence and underpin the clinical relevance of staphylococcal SigB-deficient variants which are consistently isolated during human chronic infections.
... Multiple phenotypes with decreased sensitivity to glycopeptides are on the rise due to increased vancomycin usage in critically ill patients with MRSA infections. 12 It is possible that vancomycin treatment failures against putatively vancomycin-susceptible Staphylococcus aureus are due to undiagnosed VISA or hVISA. 13 In the present study, we used both BHI-V3 and BHI-V4 screen agar for detection of hVISA and compared it with E test GRD. ...
Objectives: MRSA isolates with vancomycin MIC of 1-2 μg/ml have been linked with treatment failure and heteroresistant VISA phenotype. This study was aimed at comparing two screening methods i.e. GRD Etest and Vancomycin Screen agar in detection of heteroresistance. Material & Methods: The present study was carried out on 41 Methicillin Resistant Staphylococcus aureus strains isolated from different clinical specimens collected from Lahore General Hospital, Lahore. After screening for methicillin resistance, vancomycin MIC was determined by standard E test. Isolates with a vancomycin MIC of 1-2 μg/ml were screened for heteroresistance by Glycopeptide Resistance Detection (GRD) E-test and Vancomycin screen agar. Data was entered and analyzed by using SPSS version 20.0. Study Design: Comparative Study. Setting: Pathology Department of Post Graduate Medical Institute, Lahore. Period: May 2014 to May 2015. Results: When compared with E test GRD, Vancomycin screen agar (V3) showed 100% sensitivity with a 95% CI 39.76% to 100% and the specificity was 65 % with a 95 % CI 47.46% to 79.79%. Its PPV was 23% and NPV was 100% with an overall diagnostic accuracy of 68%. When compared with E test GRD, Vancomycin screen agar (V4) showed a sensitivity of 75% with a 95% CI 19.41% to 99.37% and a specificity of 86.47% with a 95% CI 71.91 to 95.59%. Its PPV was 37.5% and NPV predictive value was 96.96% with an overall diagnostic accuracy of 85.36%. Conclusion: In developing countries like Pakistan, where E tests are costly and difficult to use in routine laboratories, a screening test, which does not miss heteroresistant VISA may be of clinical use.
... There has been a general agreement that CC398 is increasing worldwide although information on prevalence rate has been difficult to obtain. Other complex MRSA lineages in livestock that have been found include the CC9, CC1, CC5, CC97, CC121, CC130, and ST 425 [42]. It is of interest that a human CA-MRSA type descended from bovine MSSA after bovine-host adaptation [43][44]. ...
Staphylococcus aureus is a major human pathogen associated with a variety of clinical diseases. It is the leading cause of wound infections, skin infections, respiratory infections as well as device-related infections. This review comprehensively covers the virulence determinants of the organism and the different mechanisms of antibiotic resistance in the organism. Recently, Staphylococcus aureus has become a serious threat because of its ability to evolve which has led to challenges in the treatment of infections caused by the organism.
... However, S. aureus is also an opportunistic pathogen capable of causing a variety of human diseases ranging from mild skin infections to devastating destruction of any organ system to which it gains access. S. aureus infections are a source of serious morbidity and mortality in both hospital and community settings [3,4]. ...
Purpose:
Staphylococcus aureus is an opportunistic human pathogen that can cause serious infections in humans. A plethora of known and putative virulence factors are produced by staphylococci that collectively orchestrate pathogenesis. Ear protein (Escherichia coli ampicillin resistance) in S. aureus is an exoprotein in COL strain, predicted to be a superantigen, and speculated to play roles in antibiotic resistance and virulence. The goal of this study was to determine if expression of ear is modulated by single nucleotide polymorphisms in its promoter and coding sequences and whether this gene plays roles in antibiotic resistance and virulence.
Methodology:
Promoter, coding sequences and expression of the ear gene in clinical and carriage S. aureus strains with distinct genetic backgrounds were analysed. The JE2 strain and its isogenic ear mutant were used in a systemic infection mouse model to determine the competiveness of the ear mutant.Results/Key findings. The ear gene showed a variable expression, with USA300FPR3757 showing a high-level expression compared to many of the other strains tested including some showing negligible expression. Higher expression was associated with agr type 1 but not correlated with phylogenetic relatedness of the ear gene based upon single nucleotide polymorphisms in the promoter or coding regions suggesting a complex regulation. An isogenic JE2 (USA300 background) ear mutant showed no significant difference in its growth, antibiotic susceptibility or virulence in a mouse model.
Conclusion:
Our data suggests that despite being highly expressed in a USA300 genetic background, Ear is not a significant contributor to virulence in that strain.
... HMW1, HMW2, Hia) (Duell, Su and Riesbeck 2016). Not surprisingly, some nonchemotactic pathogenic bacteria possess a range of diverse adhesins that enable them not only to colonise their preferential niches, but also to disseminate within the host by invading distant sites throughout the course of the infection (Vidal Pessolani et al. 2003;Carbonnelle et al. 2009;Speziale et al. 2009;Orrskog et al. 2012;Fitzgerald 2014;Govender, Ramsugit and Pillay 2014). ...
Chemotaxis enables microorganisms to move according to chemical gradients. Although this process requires substantial cellular energy, it also affords key physiological benefits, including enhanced access to growth substrates. Another important implication of chemotaxis is that it also plays an important role in infection and disease, as chemotaxis signaling pathways are broadly distributed across a variety of pathogenic bacteria. Furthermore, current research indicates that chemotaxis is essential for the initial stages of infection in different human, animal and plant pathogens. This review focuses on recent findings that have identified specific bacterial chemoreceptors and corresponding chemoeffectors associated with pathogenicity. Pathogenicity related chemoeffectors are either host and niche specific signals or intermediates of the host general metabolism. Plant pathogens were found to contain an elevated number of chemotaxis signaling genes and functional studies demonstrate that these genes are critical for their ability to enter the host. The expanding body of knowledge of the mechanisms underlying chemotaxis in pathogens provides a foundation for the development of new therapeutic strategies capable of blocking infection and preventing disease by interfering with chemotactic signaling pathways.
... P. aeruginosa is an opportunistic pathogen that commonly infects immunocompromised individuals, particularly in the respiratory tracts of patients with cystic fibrosis [34][35][36]. S. aureus including MRSA strains regularly establishes persistent infections of implants and post-operative wound sites [37][38][39][40]. Both P. aeruginosa and S. aureus can cause serious complications to existing medical conditions and threaten lives. ...
Statement of significance:
Nanostructured cicada and dragonfly wing surfaces have been discovered to be able physically kill bacterial cells. Here, we report on the successful fabrication of bactericidal three-dimensional structures of two main lipid components of insect wings epicuticule, palmitic (C16) and stearic (C18) fatty acids. Both palmitic and stearic acids after crystallisation on highly ordered pyrolytic graphite surfaces displayed bactericidal activity against both Gram-negative Pseudomonas aeruginosa and Gram-positive Staphylococcus aureus cells. The simplicity of the production of these microcrystallite interfaces suggests that a fabrication technique based on solution deposition could be effectively applied to form bactericidal nanocoatings.
... to form this structure (Kwon et al. 2008; Tan et al. 2012;Ross Fitzgerald 2013;Naicker et al. 2016). Moreover, the use of antibiotics may be harmful due to their incapability to distinguish S. aureus from commensal microorganisms whose reduction may facilitate the colonization by other pathogenic bacteria (Kwon et al. 2008;Ring et al. 2012). ...
Aims:
Chitosan is a natural compound that has been validated as a viable antimicrobial agent against Staphylococcus aureus. With this work we sought to evaluate the planktonic and sessile sensitivity of methicillin resistance S. aureus to chitosan's activity and evaluate if MRSA would be more or less sensitive to chitosan's activity than MSSA.
Methods and results:
A group comprised of reference strains and clinical multiresistant isolates of MSSA and MRSA were used. Methicilin resistance effect upon chitosan activity was assessed in planktonic setting and in different phases of sessile colonization, namely adhesion, biofilm formation and mature biofilm through biomass and metabolism inhibition. The results obtained showed that S. aureus methicillin resistance mechanism did not impair chitosan's activity as the highest bacterial susceptibility was registered for MRSA. Chitosan was highly effective in inhibiting MSSA and MRSA strains in both planktonic and sessile settings with biofilm inhibition percentages reaching as high as 90% for MRSA.
Conclusions:
S. aureus methicillin resistance did not impair chitosan's antimicrobial and antibiofilm activities and MRSA and MSSA were inhibited both in planktonic and sessile settings at low concentrations with great efficacy.
Significance and impact of study:
Considering the obtained results chitosan shows potential as an alternative for the control of biofilm related recalcitrant MRSA infections. This article is protected by copyright. All rights reserved.
... Colonization includes superficial residence of these bacteria on the skin as a component of the microbiome (Oh et al., 2016), intercalation within the layers of the epidermis (Nakatsuji et al., 2013), and intracellular persistence within keratinocytes (Soong et al., 2015). Skin and soft-tissue infections, as well as systemic S. aureus infection, are usually caused by colonizing strains, well documented in multiple whole-genome studies (Fitzgerald, 2014). Although typically considered to be extracellular pathogens, S. aureus can also persist within immune, endothelial, and epithelial cells that can serve as foci of persistent or recurrent infection. ...
Human skin is commonly colonized and infected by Staphylococcus aureus. Exactly how these organisms are sensed by keratinocytes has not been clearly delineated. Using a combination of metabolic and transcriptomic methodologies, we found that S. aureus infection is sensed as a metabolic stress by the hypoxic keratinocytes. This induces HIF1α signaling, which promotes IL-1β production and stimulates aerobic glycolysis to meet the metabolic requirements of infection. We demonstrate that staphylococci capable of glycolysis, including WT and agr mutants, readily induce HIF1α responses. In contrast, Δpyk glycolytic mutants fail to compete with keratinocytes for their metabolic needs. Suppression of glycolysis using 2-DG blocked keratinocyte production of IL-1β in vitro and significantly exacerbated the S. aureus cutaneous infection in a murine model. Our data suggest that S. aureus impose a metabolic stress on keratinocytes that initiates signaling necessary to promote both glycolysis and the proinflammatory response to infection.
... HMW1, HMW2, Hia) (Duell, Su and Riesbeck 2016). Not surprisingly, some nonchemotactic pathogenic bacteria possess a range of diverse adhesins that enable them not only to colonise their preferential niches, but also to disseminate within the host by invading distant sites throughout the course of the infection (Vidal Pessolani et al. 2003;Carbonnelle et al. 2009;Speziale et al. 2009;Orrskog et al. 2012;Fitzgerald 2014;Govender, Ramsugit and Pillay 2014). ...
Chemoreceptors are at the beginning of chemosensory signaling cascades that correspond to a major signal transduction mechanism. Chemoreceptors show a significant structural diversity of their ligand binding domains which present either a mono-modular or bi-modular arrangement. Although the majority of chemoreceptors are of unknown function, significant progress has been made in recent years in their functional annotation, which is reviewed here. In vitro ligand binding studies to recombinant ligand binding domains proved to be an efficient strategy to identify chemoreceptor functions. Obtained information is consistent with the view that a major driving force for the evolution of chemotaxis is to access carbon and nitrogen sources. The use of the newly generated information for the construction of biosensors is discussed.
... Staphylococcosis is common in human, animals and birds (Fitzgerald 2014, Peton et al 2014. The infection is primarily caused by Staphylococcus aureus. ...
... Ζ πνιππαξαγνληηθή αλάιπζε γηα ηνπο παξάγνληεο πνπ ζρεηίδνληαη κε ηελ παξαγσγή βηνκεκβξάλεο αλέδεημε ηελ παξνπζία ηνπ γνληδίνπ esp θαζώο θαη ζηέιερνο πνπ πξνέξρεηαη από ινίκσμε σο ζηαηηζηηθώο ζεκαληηθνύο.ΗΗΗ. Staphylococcus aureus ανθεκηικόρ ζε μεθικιλλίνη (MRSA)Ο S. aureus, θαη ζπγθεθξηκέλα ν MRSA, απνηειεί έλα ζεκαληηθό λνζνθνκεηαθό παζνγόλν παγθνζκίσο[14,257,259,[446][447][448][449][450][451][452][453][454][455][456][457][458][459][460][461][462]. Παξάγεη κία πιεζώξα ινηκνγόλσλ παξαγόλησλ πνπ ζπκβάιινπλ ζηελ παζνγέλεηα θαη δηεηζδπηηθόηεηά ηνπ, εθ ησλ νπνίσλ νη πην ζεκαληηθέο είλαη ε Panton-Valentine Leukocidin (PVL) θαη ε[500][501][502][503][504]. ...
The purpose of this study was to investigate the colonization and infections caused by KPC-producing Klebsiella pneumoniae (KPC-Kp), vancomycin-resistant Enterococcus (VRE) and methicillin-resistant Staphylococcus aureus in patients hospitalized in the Intensive Care Units of the University Hospital of Patras (ICU A) and the General Hospital “Saint Andrew” during October 2009 and February 2012.
The dissemination of KPC-Kp constitutes the most important issue in Greek ICUs, with its percentage rising in medical and surgical wards. During the duration of this study, 12.8% of patients admitted in the ICU A (52 from 405 patients) were colonized upon admission and previous ICU stay, chronic obstructive pulmonary disease, duration of previous hospitalization and previous usage of carbapenem or combination of beta-lactamic/lactamase were found to influence colonization. A gradual increase of the percentage of colonized patients admitted at the ICU from 3.9% (4 from 102 patients) during the first 6 months to 15.8% (48 from 300 patients) the next 16 months that reflects the dissemination of KPC-Kp in non-ICU wards.
Among the 226 non-colonized upon ICU A admission patients, 164 (72.6%) became colonized during their stay with the presence of colonized patients in nearby beds and the previous colonized occupant in the same bed were associated with colonization, which did not influence mortality. The high percentage of colonization in combination with the aforementioned factors indicates the importance of the dissemination of KPC-Kp among patients via the personnel and signifies the value of a strict implementation of infection control protocols.
In total, 53 patients developed KPC-Kp bloodstream infection during ICU A stay with 43.4% mortality. The most important factors that influence mortality were the resistance of the strain to gentamicin/colistin/tigecycline and septic shock, while the treatment with two active antibiotics was associated with better survival confirming the results of previous studies favoring combination therapy for the treatment of KPC-Kp infection.
The development of resistance against colistin or tigecycline, which are considered the last frontier in the treatment of KPC-Kp infections, is an alarming phenomenon. In total, 24.4% and 17.9% of ICU A patients became colonized by KPC-Kp resistant to colictin or tigecycline, respectively. As expected, the administration of colistin or tigecycline influenced colonization, while the most important factor favoring colonization was the presence of colonized patients in nearby patients, indicating the importance of dissemination of these strains against de novo resistance development.
The comparison of the two ICUs, found a higher percentage of patients colonized during ICU A stay (61.8% vs 34.1%) and in a shorter period (10.6 vs 19.9 days). These results may be explained by the higher percentage of patients colonized upon admission (11.4% vs 1.8%), the lower nurse/patient ration and the higher carbapenem administration.
In total, 305 and 100 strains of K. pneumoniae isolated from patients hospitalized in ICU A and B, respectively, were positive for the presence of blaKPC gene while five strains in ICU A were positive for the blaVIM gene also. All strains were resistant to penicillins, cephalosporins, aztreonam, trimethoprim sulfamethoxazole (30% of ICU B strains were sensitive), amikacin, tombramycin and quinolones. The resistance rates to carbapenems (67.9% vs 60%), colisitn (35.1% vs 18%), gentamicin (50.8% vs 24%) and tigecycline (17% vs 18%) among the ICUs strains were comparable. PFGE of 57 and 20 isolates from ICU A and B, respectively, revealed that ICU A strains belonged in two types, with type A comprising 65.5% of the isolates, while all ICU B isolates belonged in type A.
The percentage of VRE colonization in both ICUs were lower in comparison with those of KPC-Kp. During ICU admission 14.3% (71 from 497 patients) was already colonized, while 14.4% (36 from 250 patients) became colonized during stay. The most important factor influencing colonization was the presence of colonized patients in nearby beds, indicating that non adherence with hand hygiene may play a predominate role in VRE dissemination.
In total 107 VRE strains were isolated (100 E. faecium and 7 E. faecalis). Eighty four were positive for the vanA gene and resistant to vancomycin and teicoplanin, while the rest were vanB positive and were characterized by low level resistance to vancomycin (12 were in susceptibility range) and susceptible to teicoplanin. All strains were susceptible to linezolid, daptomycin and tigecycline. As MLST revealed, E. faecium strains belonged in six different Sequencing Types (ST117, ST17, ST203, ST226, ST786, ST125) with 90% among them belonging to the Clonal Complex CC17. E. faecalis strains were categorized in four STs (ST6, ST41, ST19, ST28).
The proportion of colonized patients by MRSA upon admission and during ICU stay was very low (5.3% and 3.7%, respectively). The most important factor associated with colonization was enteric carriage of vanA-positive Enterococcus. Surveillance cultures revealed 28 mecA-positive S. aureus strains, with the majority (n=19) being PVL-positive, belonging to ST80 and resistant only to kanamycin, tetracycline and fucidic acid, while the remaining were categorized in four STs (6 strains in ST239 and one at ST225, ST72 and ST30). The ST30 strain was tst-positive.
The comparison of colonization strains from patients (n=67) and personnel (n=23) of the ICUs (Group A) with the strains of colonization (n=53) and bloodstream infections (n=75) isolated from non-ICU patients (Group B), revealed a higher percentage of MRSA and PVL-positive strains in Group B, while Group A was characterized by higher percentage of tst-positive strains indicating their silent dissemination between ICU patients and personnel.
The present study has identified the risk factors for colonization of infection by KPC-Kp, VRE and MRSA, in order to guide the future efforts towards containing their dissemination in the two ICUs, as well as, to the Greek hospitals, which in total are plagued by the aforementioned pathogens.
... Many of these same bacterial traits are also known to evolve rapidly in clinical infections and this evolution may in some cases contribute to important changes in patient prognosis or the response of the infection to treatment (e.g. Fitzgerald, 2014;Folkesson et al., 2012). The evolutionary dynamics of chronic bacterial lung infections associated with cystic fibrosis (CF) have been particularly well-described . ...
It is increasingly clear that rapid evolutionary dynamics are an important process in microbial ecology. Experimental evolution, wherein microbial evolution is observed in real-time, has revealed many instances of appreciable evolutionary change occurring on very short timescales of a few days or weeks in response to a variety of biotic and abiotic selection pressures. From clinical infections, including the chronic bacterial lung infections associated with cystic fibrosis that form a focus of my research, there is now abundant evidence suggesting that rapid evolution by infecting microbes contributes to host adaptation, treatment failure and worsening patient prognosis. However, disentangling the drivers of natural selection in complex infection environments is extremely challenging and limits our understanding of the selective pressures acting upon microbes in infections. Controlled evolution experiments can make a vital contribution to this by determining the causal links between predicted drivers of natural selection and the evolutionary responses of microbes. Integration of experimental evolution into studies of clinical infections is a key next step towards a better understanding of the causes and consequences of rapid microbial evolution in infections and discovering how these evolutionary processes might be influenced to improve patient health.
... In recent years, the incidence of S. aureus-and P. aeruginosa-linked infections has increased, with highly virulent strains being encountered. In addition, the ability of Staphylococcus and Pseudomonas species to survive on environmental surfaces increases the probability of persistent contamination in household settings [5] . The aforementioned pathogens can be acquired from hospitals and also increasingly from non-clinical environments (i.e., community-associated infections). ...
A study was performed to estimate the prevalence of the external bacterial flora of two domestic cockroaches (Blattella germanica and Blatta orientalis) collected from households in Tebessa (northeast Algeria). Three major bacterial groups were cultured (total aerobic, enterobacteria, and staphylococci) from 14 specimens of cockroaches, and antibiotic susceptibility was tested for both Staphylococcus and Pseudomonas isolates. Culturing showed that the total bacterial load of cockroaches from different households were comparable (P<0.001) and enterobacteria were the predominant colonizers of the insect surface, with a bacterial load of (2.1×105 CFU/insect), whereas the staphylococci group was the minority. Twenty-eight bacterial species were isolated, and susceptibility patterns showed that most of the staphylococci isolates were highly susceptible to chloramphenicol, gentamycin, pristinamycin, ofloxacin, clindamycin, and vancomycin; however, Pseudomonas strains exhibited resistance to amoxicillin/clavulanic acid, imipenem, and the second-generation antibiotic cephalosporin cefuroxime.
Copyright © 2015 The Editorial Board of Biomedical and Environmental Sciences. Published by China CDC. All rights reserved.
Skin microbiota is an integral part of the human immune system. Staphylococcus aureus is one of the essential components of the normal flora. Approximately 20-30% of healthy individuals are persistently colonized with S. aureus, whereas the remainders are considered low-level intermittent carriers. Despite these natural aspects of existence, S. aureus can be a major opportunistic human pathogen. This versatile microorganism can infect a variety of anatomical sites, causing a broad spectrum of pathologies ranging from superficial to invasive infections. It developed a variety of strategies to adopt to a changing microenvironment. This attributed to the emergence of resistance to antibiotics of different classes during the past six decades. Methicillin-resistant S. aureus (MRSA) was originally confined to health-care settings (health-care-associated MRSA). Later on, community-acquired MRSA was identified as another source of infections. Recent figures indicate that MRSA strains have been associated with approximately 75% of all S. aureus infections worldwide. Several guidelines have been published to establish an adequate treatment of skin and soft tissue infections (SSTIs) caused by MRSA strains. In the first part of this review, we focus on current treatment guidelines with a focus on medical drug therapy, but drug therapy has its own limitations. Recently, the interest in herbal remedies has greatly increased. There is growing evidence of antimicrobial activity of medicinal plants and their extracts. The second part of this review is dedicated to herbal compounds to circumvent antibiotic resistance. Herbal compounds may potentiate the action of antibiotics and restore the activities of antibacterial agents against which S. aureus has developed a drug resistance. Part 2 focuses on the role of S. aureus in pathology of the two major inflammatory skin diseases, i.e., atopic dermatitis (AD) and psoriasis. Finally, Part 3 provides an overview on natural compounds with antimicrobial activity against S. aureus and possible use in the treatment of SSTIs,
Staphylococcus aureus is an important human pathogen but is primarily a commensal of the human nose and skin. Survival during colonisation is likely one of the major drivers of S. aureus evolution. Here we use a genome-wide mutation enrichment approach to analyse a genomic dataset of 3,060 S. aureus isolates from 791 individuals to show that despite limited within-host genetic diversity, an excess of protein-altering mutations can be found in genes encoding key metabolic pathways, in regulators of quorum-sensing and in known antibiotic targets. Nitrogen metabolism and riboflavin synthesis are the metabolic processes with strongest evidence of adaptation. Further evidence of adaptation to nitrogen availability was revealed by enrichment of mutations in the assimilatory nitrite reductase and urease, including mutations that enhance growth with urea as the sole nitrogen source. Inclusion of an additional 4,090 genomes from 802 individuals revealed eight additional genes including sasA/sraP, pstA, and rsbU with signals adaptive variation that warrant further characterisation. Our study provides the most comprehensive picture to date of the heterogeneity of adaptive changes that occur in the genomes of S. aureus during colonisation, revealing the likely importance of nitrogen metabolism, loss of quorum sensing and antibiotic resistance for successful human colonisation.
This study was performed to examine molecular docking of newly synthesised thiazolidinone compounds and discover potential InhA inhibitors. MIC screening produced thiazolidinone derivatives for Staphylococcus aureus illness. KBr pellet FT-IR spectra were collected on a Thermo Nicolet AVATAR-330 spectrometer. Bruker obtained 400 MHz 1H spectra of all chemicals in DMSO-d6. Five 4-thiazolidinone derivatives:4-(4-oxo-2-phenylthiazolidin-3yl)amino)benzyl)oxazolidin-2-one(8), ((2-(4-chlorophenyl)-4oxothiazolidin-3yl)amino)benzyl)-oxazolidin-2-one(9), ((2-(4-fluorophenyl)-4-oxothiazolidin-3-yl)amino)benzyl)oxazolidin-2-one(10), 4-(4-((4-oxo-2-(p-tolyl)thiazolidin-3yl)amino)benzyl)oxazolidin-2-one(11) and 4-(4-((2-(4-methoxyphenyl)-4-oxothiazolidin-3yl)amino)benzyl)oxazolidin-2-one(12) docked due to its vast biological usefulness. Molecular docking, ADMET, TOPKAT Toxicity, and molecular dynamic simulation were used to estimate drug-likeness for 1QG6. MIC screening of synthesised thiazolidinone derivatives against Staphylococcus aureus through disc diffusion. Molecular docking studies examining the inhibitory effect of produced substances suggest testing for MIC antibacterial activity. Compound 12's antibacterial activity against Staphylococcus aureus is compared to Ampicillin and Cefixime. Compound 12 is utilised as an alternate medication for Staphylococcus aureus. A.: Identification of potential inhibitor targeting InhA, molecular docking, ADMET, molecular dynamic simulation and antibacterial activity of thiazolidinone derivatives: A computational approach. Intern. J. Zool. Invest. 8(Special Issue):191-202, 2022.
Espécies do gênero Staphylococcus estão entre os principais agentes causadores de infecções de origem nosocomial, e é especialmente preocupante o aumento de cepas multidrogarresistentes que circulam no ambiente hospitalar, colonizando e infectando pacientes e profissionais de saúde. Infecções secundárias são um agravo importante na COVID-19. O objetivo desse trabalho é analisar através da literatura científica atual o impacto da infecção concomitante por espécies do gênero Staphylococcus em pacientes com COVID-19. A análise da literatura científica atual permite inferir que há uma grande heterogeneidade quanto à incidência dessa coinfecção/superinfecção/colonização. Também foi verificada heterogeneidade em relação aos níveis de resistência a antibióticos das cepas isoladas e tendências divergentes quanto à incidência temporal desse patógeno bacteriano nas infecções secundárias à COVID-19. Essas diferenças provavelmente refletem padrões locais, como os perfis de resistência e infecções secundárias anteriores à pandemia, assim como os níveis de controle de infecções hospitalares de cada unidade de saúde. Fatores como o tempo de internação, o uso de ventilação mecânica e a aquisição de cepas de Staphylococcus spp. de origem hospitalar influenciam diretamente nas taxas de infecção secundária e o prognóstico em pacientes com a COVID-19. Mesmo considerando a heterogeneidade dos resultados, as taxas de mortalidade podem ser consideradas altas em pacientes que apresentam infecção concorrente por Staphylococcus spp. na forma grave da virose causada pelo SARS-CoV-2, com taxas que podem ser superiores a 90%.
Detection of pathogenic bacteria in complex biological matrices remains a major challenge. Herein, we report the selection and optimization of a new DNAzyme for Staphylococcus aureus (SA) and the use of the DNAzyme to develop a simple lateral flow device (LFD) for detection of SA in nasal mucus. The DNAzyme was generated by in vitro selection using a crude extra/intracellular mixture derived from SA, which could be used directly for simple solution or paper-based fluorescence assays for SA. The DNAzyme was further modified to produce a DNA cleavage fragment that acted as a bridging element to bind DNA-modified gold nanoparticles to the test line of a LFD, producing a simple colorimetric dipstick test. The LFD was evaluated with nasal mucus samples spiked with SA, and demonstrated that SA detection was possible in minutes with minimal sample processing.
Detection of pathogenic bacteria in complex biological matrices remains a major challenge. Herein, we report the selection and optimization of a new DNAzyme for Staphylococcus aureus (SA) and the use of the DNAzyme to develop a simple lateral flow device (LFD) for detection of SA in nasal mucus. The DNAzyme was generated by in vitro selection using a crude extra/intracellular mixture derived from SA, which could be used directly for simple solution or paper‐based fluorescence assays for SA. The DNAzyme was further modified to produce a DNA cleavage fragment that acted as a bridging element to bind DNA‐modified gold nanoparticles to the test line of a LFD, producing a simple colorimetric dipstick test. The LFD was evaluated with nasal mucus samples spiked with SA, and demonstrated that SA detection was possible in minutes with minimal sample processing.
Adaptation of Staphylococcus aureus to host microenvironments during chronic infection involves spontaneous mutations, yet changes underlying adaptive phenotypes remain incompletely explored. Here we employed artificial selection and whole genome sequencing to better characterize spontaneous chromosomal mutations that alter two pathogenicity phenotypes relevant to chronic infection in S. aureus : intracellular invasiveness and intracellular cytotoxicity. We identified 23 genes whose alteration coincided with enhanced virulence, 11 that were previously known and 12 of which (52%) had no previously described role in S. aureus pathogenicity. Using precision genome editing, transposon mutants, and gene complementation, we empirically assessed the contributions of individual genes to the two virulence phenotypes. We functionally validated 14 of 21 genes tested as measurably influencing invasion and/or cytotoxicity, including 8 newly implicated by this study. We identified inactivating mutations ( murA , ndhC , and a hypothetical membrane protein) and gain-of-function mutations ( aroE Thr182Ile, yhcF Thr74Ile, and Asp486Glu in a hypothetical peptidase) in previously unrecognized S. aureus virulence genes that enhance pathogenesis when introduced into a clean genetic background, as well as a novel activating mutation in known virulence regulator saeS (Ala106Thr). Investigation of potentially epistatic interactions identified a tufA mutation (Ala271Val) that enhances virulence only in the context of purine operon repressor ( purR ) inactivation. This project reveals a functionally diverse range of genes affected by gain- or loss-of-function mutations that contribute to S. aureus adaptive virulence phenotypes. More generally, the work establishes artificial selection as a means to determine the genetic mechanisms underlying complex bacterial phenotypes relevant to adaptation during infection.
The bacterial pathway for fatty acid biosynthesis, FASII, is a target for development of new anti-staphylococcal drugs. This strategy is based on previous reports indicating that self-synthesized fatty acids appear to be indispensable for Staphylococcus aureus growth and virulence, although other bacteria can use exogenous fatty acids to compensate FASII inhibition. Here we report that staphylococci can become resistant to the FASII-targeted inhibitor triclosan via high frequency mutations in fabD, one of the FASII genes. The fabD mutants can be conditional for FASII and not require exogenous fatty acids for normal growth, and can use diverse fatty acid combinations (including host fatty acids) when FASII is blocked. These mutants show cross-resistance to inhibitors of other FASII enzymes and are infectious in mice. Clinical isolates bearing fabD polymorphisms also bypass FASII inhibition. We propose that fatty acid-rich environments within the host, in the presence of FASII inhibitors, might favour the emergence of staphylococcal strains displaying resistance to multiple FASII inhibitors.
Evaluation of the antimicrobial efficiency of antibiotic-polymer association complexes on different strains of Staphylococcus aureus
The virulence of Staphylococcus aureus, in both human and animal hosts, is largely influenced by the acquisition of mobile genetic elements (MGEs). Most S. aureus strains carry a variety of MGEs, including three genomic islands (νSaα, νSaβ, νSaγ) that are diverse in virulence gene content but conserved within strain lineages. Although the mobilization of pathogenicity islands, phages and plasmids has been well studied, the mobilization of genomic islands is poorly understood. We previously demonstrated the mobilization of νSaβ by the adjacent temperate bacteriophage ϕSaBov from strain RF122. In this study, we demonstrate that ϕSaBov mediates the mobilization of νSaα and νSaγ, which are located remotely from ϕSaBov, mostly to recipient strains belonging to ST151. Phage DNA sequence analysis revealed that chromosomal DNA excision events from RF122 were highly specific to MGEs, suggesting sequence-specific DNA excision and packaging events rather than generalized transduction by a temperate phage. Disruption of the int gene in ϕSaBov did not affect phage DNA excision, packaging, and integration events. However, disruption of the terL gene completely abolished phage DNA packing events, suggesting that the primary function of temperate phage in the transfer of genomic islands is to allow for phage DNA packaging by TerL and that transducing phage particles are the actual vehicle for transfer. These results extend our understanding of the important role of bacteriophage in the horizontal transfer and evolution of genomic islands in S. aureus.
A key to persistent and recurrent Staphylococcus aureus infections is its ability to adapt to diverse and toxic conditions. This ability includes a switch into a biofilm or to the quasi-dormant Small Colony Variant (SCV). The development and molecular attributes of SCVs have been difficult to study due to their rapid reversion to their parental cell-type. We recently described the unique induction of a matrix-embedded and stable SCV cell-type in a clinical S. aureus strain (WCH-SK2) by growing the cells with limiting conditions for a prolonged timeframe. Here we further study their characteristics. They possessed an increased viability in the presence of antibiotics compared to their non-SCV form. Their stability implied there had been genetic changes, we therefore determined both the genome sequence of WCH-SK2 and its stable SCV form at a single base resolution, employing Single Molecular Real-Time (SMRT) sequencing that enabled the methylome to also be determined. The genetic features of WCH-SK2 have been identified; the SCCmec type, the pathogenicity and genetic islands and virulence factors. The genetic changes that had occurred in the stable SCV form were identified; most notably being in MgrA, a global regulator, and RsbU, a phosphoserine phosphatase within the regulatory pathway of the sigma factor SigB. There was a shift in the methylomes of the non-SCV and stable SCV forms. We have also shown a similar induction of this cell-type in other S. aureus strains and performed a genetic comparison to these and other S. aureus genomes. We additionally map RNAseq data to the WCH-SK2 genome in a transcriptomic analysis of the parental, SCV and stable SCV cells. The results from this study represent the unique identification of a suite of epigenetic, genetic and transcriptional factors that are implicated in the switch in S. aureus to its persistent SCV form.
Gastrointestinal diseases have a huge impact especially in third world countries, making it urgent to seek new effective antimicrobial therapies. Thus, the development of nanoparticles (NPs) with bioactive compounds having antimicrobial activity has been the target of research over the past years. The development of antimicrobial drug NPs may be promising to overcome the problems associated with antibiotic resistance caused by many pathogenic bacteria. Moreover, the NPs administration of antimicrobial agents has advantages associated therewith, as use of low cost materials, contribution to the improvement of the therapeutic index and a controlled release drug by increasing the pharmacokinetics. These systems can be used to specific strains of bacteria, and to release interesting antimicrobial compounds. The phenolic compounds (PC) are a class of such bioactive compounds for which their antimicrobial activity was already tested on the production of NPs. Polymeric or lipidic NPs systems have been investigated to deliver these compounds. Chitosan is a polymer widely known for their properties, especially the antimicrobial activity and its ability to adhere to intestinal epithelium. This review article aims to evaluate and discuss recent developments in PC new delivery systems with antimicrobial activity against gastrointestinal pathogens, their production processes, activities, focusing on NPs produced using chitosan as the main structural and functional material.
Copyright © 2015 Elsevier Ltd. All rights reserved.
To compare the in vitro antimicrobial efficacy of ceftaroline with linezolid against Staphylococcus aureus and methicillin resistant Staphylococcus aureus.
Quasi-experimental study.
Microbiology Department, Army Medical College, Rawalpindi, from January to December 2013.
Clinical samples from respiratory tract, blood, pus and various catheter tips routinely received in the Department of Microbiology, Army Medical College, Rawalpindi were innoculated on blood and MacConkey agar. Staphylococcus aureus was identified by colony morphology, Gram reaction, catalase test and coagulase test. Methicillin resistant Staphylococcus aureus detection was done by modified Kirby Bauer disc diffusion method using cefoxitin disc (30μg) and the isolates were considered methicillin resistant if the zone of inhibition around cefoxitin disc was ² 21 mm. Bacterial suspensions of 56 Staphylococcus aureus isolates and 50 MRSA isolates were prepared, which were standardized equal to 0.5 McFarland's turbidity standard and inoculated on Mueller-Hinton agar plates followed by application of ceftaroline and linezolid disc (Oxoid, UK), according to manufacturer's instructions. The plates were then incubated at 37°C aerobically for 18 - 24 hours. Diameters of inhibition zone were measured and interpretated as per Clinical and Laboratory Standards Institute (CLSI) guidelines.
Out of 106 isolates all of the 56 Staphylococcus aureus (100%) were sensitive to ceftaroline and linezolid. However, out of 50 methicillin resistant Staphylococcus aureus, 48 (96%) were sensitive to ceftaroline whereas, 49 (98%) were sensitive to linezolid.
Ceftaroline is equally effective as linezolid against Staphylococcus aureus and methicillin resistant Staphylococcus aureus.
Staphylococcus aureus is a major pathogen of humans and animals. The capacity of S. aureus to adapt to different host species and tissue types is strongly influenced by the acquisition of mobile genetic elements encoding determinants involved in niche adaptation. The genomic islands νSaα and νSaβ are found in almost all S. aureus strains and are characterized by extensive variation in virulence gene content. However the basis for the diversity and the mechanism underlying mobilization of the genomic islands between strains are unexplained. Here, we demonstrated that the genomic island, νSaβ, encoding an array of virulence factors including staphylococcal superantigens, proteases, and leukotoxins, in addition to bacteriocins, was transferrable in vitro to human and animal strains of multiple S. aureus clones via a resident prophage. The transfer of the νSaβ appears to have been accomplished by multiple conversions of transducing phage particles carrying overlapping segments of the νSaβ. Our findings solve a long-standing mystery regarding the diversification and spread of the genomic island νSaβ, highlighting the central role of bacteriophages in the pathogenic evolution of S. aureus.
The widespread use of antibiotics in association with high-density clinical care has driven the emergence of drugresistant bacteria that are adapted to thrive in hospitalized patients. Of particular concern are globally disseminated methicillin-resistant Staphylococcus aureus (MRSA) clones that cause outbreaks and epidemics associated with health care. The most rapidly spreading and tenacious health-care-associated clone in Europe currently is EMRSA-15, which was first detected in the UK in the early 1990s and subsequently spread throughout Europe and beyond. Using phylogenomic methods to analyze the genome sequences for 193 S. aureus isolates, we were able to show that the current pandemic population of EMRSA-15 descends from a health-care-associatedMRSA epidemic that spread throughout England in the 1980s, which had itself previously emerged froma primarily community-associated methicillin-sensitive population. The emergence of fluoroquinolone resistance in this EMRSA-15 subclone in the English Midlands during the mid-1980s appears to have played a key role in triggering pandemic spread, and occurred shortly after the first clinical trials of this drug. Genome-based coalescence analysis estimated that the population of this subclone over the last 20 yr has grown four times faster than its progenitor. Using comparative genomic analysis we identified the molecular genetic basis of 99.8% of the antimicrobial resistance phenotypes of the isolates, highlighting the potential of pathogen genome sequencing as a diagnostic tool.We document the genetic changes associated with adaptation to the hospital environment and with increasing drug resistance over time, and how MRSA evolution likely has been influenced by country-specific drug use regimens.
Unlabelled:
Staphylococcus aureus is a human commensal that at times turns into a serious bacterial pathogen causing life-threatening infections. For the delicate control of virulence, S. aureus employs the agr quorum-sensing system that, via the intracellular effector molecule RNAIII, regulates virulence gene expression. We demonstrate that the presence of the agr locus imposes a fitness cost on S. aureus that is mediated by the expression of RNAIII. Further, we show that exposure to sublethal levels of the antibiotics ciprofloxacin, mupirocin, and rifampin, each targeting separate cellular functions, markedly increases the agr-mediated fitness cost by inducing the expression of RNAIII. Thus, the extensive use of antibiotics in hospitals may explain why agr-negative variants are frequently isolated from hospital-acquired S. aureus infections but rarely found among community-acquired S. aureus strains. Importantly, agr deficiency correlates with increased duration of and mortality due to bacteremia during antibiotic treatment and with a higher frequency of glycopeptide resistance than in agr-carrying strains. Our results provide an explanation for the frequent isolation of agr-defective strains from hospital-acquired S. aureus infections and suggest that the adaptability of S. aureus to antibiotics involves the agr locus.
Importance:
Staphylococcus aureus is the most frequently isolated pathogen in intensive care units and a common cause of nosocomial infections, resulting in a high degree of morbidity and mortality. Surprisingly, a large fraction (15 to 60%) of hospital-isolated S. aureus strains are agr defective and lack the main quorum-sensing-controlled virulence regulatory system. This is a problem, as agr-defective strains are associated with a mortality level in bacteremic infections and a probability of glycopeptide resistance greater than those of other strains. We show here that agr-negative strains have a fitness advantage over agr-positive strains in the presence of sublethal concentrations of some antibiotics and that the fitness defect of agr-positive cells is caused by antibiotic-mediated expression of the agr effector molecule RNAIII. These results offer an explanation of the frequent isolation of agr-defective S. aureus strains in hospitals and will influence how we treat S. aureus infections.
The pathogen Staphylococcus aureus undergoes phenotype switching in vivo from its normal colony phenotype (NCP) to a slow-growing, antibiotic-resistant small-colony-variant (SCV) phenotype that
is associated with persistence in host cells and tissues. However, it is not clear whether phenotype switching is the result
of a constitutive process that is selected for under certain conditions or is triggered by particular environmental stimuli.
Examination of cultures of diverse S. aureus strains in the absence of selective pressure consistently revealed a small gentamicin-resistant SCV subpopulation that emerged
during exponential-phase NCP growth and increased in number until NCP stationary phase. Treatment of replicating bacteria
with the antibiotic gentamicin, which inhibited NCP but not SCV replication, resulted in an initial decrease in SCV numbers,
demonstrating that SCVs arise as a consequence of NCP replication. However, SCV population expansion in the presence of gentamicin
was reestablished by selection of phenotype-stable SCVs and subsequent SCV replication. In the absence of selective pressure,
however, phenotype switching was bidirectional and occurred at a high frequency during NCP replication, resulting in SCV turnover.
In summary, these data demonstrate that S. aureus phenotype switching occurs via a constitutive mechanism that generates a dynamic, antibiotic-resistant subpopulation of bacteria
that can revert to the parental phenotype. The emergence of SCVs can therefore be considered a normal part of the S. aureus life cycle and provides an insurance policy against exposure to antibiotics that would otherwise eliminate the entire population.
Genome inversions are ubiquitous in organisms ranging from prokaryotes to eukaryotes. Typical examples can be identified by comparing the genomes of two or more closely related organisms, where genome inversion footprints are clearly visible. Although the evolutionary implications of this phenomenon are huge, little is known about the function and biological meaning of this process. Here, we report our findings on a bacterium that generates a reversible, large-scale inversion of its chromosome (about half of its total genome) at high frequencies of up to once every four generations. This inversion switches on or off bacterial phenotypes, including colony morphology, antibiotic susceptibility, hemolytic activity, and expression of dozens of genes. Quantitative measurements and mathematical analyses indicate that this reversible switching is stochastic but self-organized so as to maintain two forms of stable cell populations (i.e., small colony variant, normal colony variant) as a bet-hedging strategy. Thus, this heritable and reversible genome fluctuation seems to govern the bacterial life cycle; it has a profound impact on the course and outcomes of bacterial infections.
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Methicillin-resistant Staphylococcus aureus (MRSA) strains are leading causes of hospital-acquired infections in the United States, and clonal cluster 5 (CC5) is the predominant lineage responsible for these infections. Since 2002, there have been 12 cases of vancomycin-resistant S. aureus (VRSA) infection in the United States-all CC5 strains. To understand this genetic background and what distinguishes it from other lineages, we generated and analyzed high-quality draft genome sequences for all available VRSA strains. Sequence comparisons show unambiguously that each strain independently acquired Tn1546 and that all VRSA strains last shared a common ancestor over 50 years ago, well before the occurrence of vancomycin resistance in this species. In contrast to existing hypotheses on what predisposes this lineage to acquire Tn1546, the barrier posed by restriction systems appears to be intact in most VRSA strains. However, VRSA (and other CC5) strains were found to possess a constellation of traits that appears to be optimized for proliferation in precisely the types of polymicrobic infection where transfer could occur. They lack a bacteriocin operon that would be predicted to limit the occurrence of non-CC5 strains in mixed infection and harbor a cluster of unique superantigens and lipoproteins to confound host immunity. A frameshift in dprA, which in other microbes influences uptake of foreign DNA, may also make this lineage conducive to foreign DNA acquisition.
Importance:
Invasive methicillin-resistant Staphylococcus aureus (MRSA) infection now ranks among the leading causes of death in the United States. Vancomycin is a key last-line bactericidal drug for treating these infections. However, since 2002, vancomycin resistance has entered this species. Of the now 12 cases of vancomycin-resistant S. aureus (VRSA), each was believed to represent a new acquisition of the vancomycin-resistant transposon Tn1546 from enterococcal donors. All acquisitions of Tn1546 so far have occurred in MRSA strains of the clonal cluster 5 genetic background, the most common hospital lineage causing hospital-acquired MRSA infection. To understand the nature of these strains, we determined and examined the nucleotide sequences of the genomes of all available VRSA. Genome comparison identified candidate features that position strains of this lineage well for acquiring resistance to antibiotics in mixed infection.
Whole-genome sequencing offers new insights into the evolution of bacterial pathogens and the etiology of bacterial disease. Staphylococcus aureus is a major cause of bacteria-associated mortality and invasive disease and is carried asymptomatically by 27% of adults. Eighty percent of bacteremias match the carried strain. However, the role of evolutionary change in the pathogen during the progression from carriage to disease is incompletely understood. Here we use high-throughput genome sequencing to discover the genetic changes that accompany the transition from nasal carriage to fatal bloodstream infection in an individual colonized with methicillin-sensitive S. aureus. We found a single, cohesive population exhibiting a repertoire of 30 single-nucleotide polymorphisms and four insertion/deletion variants. Mutations accumulated at a steady rate over a 13-mo period, except for a cluster of mutations preceding the transition to disease. Although bloodstream bacteria differed by just eight mutations from the original nasally carried bacteria, half of those mutations caused truncation of proteins, including a premature stop codon in an AraC-family transcriptional regulator that has been implicated in pathogenicity. Comparison with evolution in two asymptomatic carriers supported the conclusion that clusters of protein-truncating mutations are highly unusual. Our results demonstrate that bacterial diversity in vivo is limited but nonetheless detectable by whole-genome sequencing, enabling the study of evolutionary dynamics within the host. Regulatory or structural changes that occur during carriage may be functionally important for pathogenesis; therefore identifying those changes is a crucial step in understanding the biological causes of invasive bacterial disease.
Antimicrobial resistance in Staphylococcus aureus is a major public health threat, compounded by emergence of strains with resistance to vancomycin and daptomycin, both last line antimicrobials. Here we have performed high throughput DNA sequencing and comparative genomics for five clinical pairs of vancomycin-susceptible (VSSA) and vancomycin-intermediate ST239 S. aureus (VISA); each pair isolated before and after vancomycin treatment failure. These comparisons revealed a frequent pattern of mutation among the VISA strains within the essential walKR two-component regulatory locus involved in control of cell wall metabolism. We then conducted bi-directional allelic exchange experiments in our clinical VSSA and VISA strains and showed that single nucleotide substitutions within either walK or walR lead to co-resistance to vancomycin and daptomycin, and caused the typical cell wall thickening observed in resistant clinical isolates. Ion Torrent genome sequencing confirmed no additional regulatory mutations had been introduced into either the walR or walK VISA mutants during the allelic exchange process. However, two potential compensatory mutations were detected within putative transport genes for the walK mutant. The minimal genetic changes in either walK or walR also attenuated virulence, reduced biofilm formation, and led to consistent transcriptional changes that suggest an important role for this regulator in control of central metabolism. This study highlights the dramatic impacts of single mutations that arise during persistent S. aureus infections and demonstrates the role played by walKR to increase drug resistance, control metabolism and alter the virulence potential of this pathogen.
The molecular adaptation of Staphylococcus aureus to its host during chronic infection is not well understood. Comparative genome sequencing of 3 S. aureus isolates obtained sequentially over 26 months from the airways of a cystic fibrosis patient, revealed variation in phage content, and genetic polymorphisms in genes which influence antibiotic resistance, and global regulation of virulence. The majority of polymorphisms were isolate-specific suggesting the existence of an heterogeneous infecting population that evolved from a single infecting strain of S. aureus. The genetic variation identified correlated with differences in growth rate, hemolytic activity, and antibiotic sensitivity, implying a profound effect on the ecology of S. aureus. In particular, a high frequency of mutations in loci associated with the alternate transcription factor SigB, were observed. The identification of genes under diversifying selection during long-term infection may inform the design of novel therapeutics for the control of refractory chronic infections.
Using multinational collections of methicillin-susceptible Staphylococcus aureus (MSSA) isolates from infective endocarditis (IE) and soft tissue infections (STIs), we sought to (1) validate the finding that S. aureus in clonal complex (CC) 30 is associated with hematogenous complications and (2) test the hypothesis that specific genetic characteristics in S. aureus are associated with infection severity.
IE and STI isolates from 2 cohorts were frequency matched by geographic origin. Isolates underwent spa typing to infer CC and multiplex polymerase chain reaction for presence of virulence genes.
114 isolate pairs were genotyped. IE isolates were more likely to be CC30 (19.5% vs 6.2%; P = .005) and to contain 3 adhesins (clfB, cna, map/eap; P < .0001 for all) and 5 enterotoxins (tst, sea, sed, see, and sei; P ≤ .005 for all). CC30 isolates were more likely to contain cna, tst, sea, see, seg, and chp (P < .05 for all).
MSSA IE isolates were significantly more likely to be CC30 and to possess a distinct repertoire of virulence genes than MSSA STI isolates from the same region. The genetic basis of this association requires further study.
Methicillin-resistant Staphylococcus aureus (MRSA), once restricted to hospitals, is spreading rapidly through the wider community. Resistance to vancomycin, the principal
drug used to treat MRSA infections, has only recently emerged, is mainly low level, and characteristically appears during
vancomycin therapy (vancomycin-intermediate S. aureus [VISA] and hetero-resistant VISA). This phenomenon suggests the adaptation of MRSA through mutation, although defining the
mutations leading to resistance in clinical isolates has been difficult. We studied a vancomycin-susceptible clinical MRSA
isolate (MIC of 1 μg/ml) and compared it with an isogenic blood culture isolate from the same patient, despite 42 days of
vancomycin treatment (MIC of 4 μg/ml). A whole-genome sequencing approach allowed the nearly complete assembly of the genome
sequences of the two isolates and revealed only six nucleotide substitutions in the VISA strain compared with the parent strain.
One mutation occurred in graS, encoding a putative two-component regulatory sensor, leading to a change from a polar to a nonpolar amino acid (T136I) in
the conserved histidine region of the predicted protein. Replacing the graS allele of the vancomycin-susceptible parent strain with the graS allele from the VISA derivative resulted in increased vancomycin resistance at a level between those of the vancomycin-susceptible
S. aureus and VISA clinical isolates, confirming a role for graRS in VISA. Our study suggests that MRSA is able to develop clinically significant vancomycin resistance via a single point
mutation, and the two-component regulatory system graRS is a key mediator of this resistance. However, additional mutations are likely required to express the full VISA phenotype.
In this study we characterized a genetic locus that is predicted to encode one of the three AraC-like regulators of Neisseria meningitidis, a homologue of MpeR of Neisseria gonorrhoeae which is specific to the pathogenic Neisseria species. Previous microarray studies have suggested that this gene is a member of the Fur regulon. In strain MC58, it is a pseudogene (annotated as two ORFs, NMB1879 and NMB1878) containing a frameshift mutation which we show is common to all strains tested belonging to the ST-32 hypervirulent clonal complex. Using primer extension and S1 nuclease protection assays, we mapped two promoters in the upstream intergenic region: the mpeR promoter and the NMB1880 promoter. The latter promoter drives transcription of the divergent upstream locus, which is predicted to encode a high-affinity iron uptake system. We demonstrated that both promoters are induced during iron limitation and that this regulation is also mediated by the Fur regulator. DNA-binding studies with the purified MpeR protein revealed that it binds to a region directly upstream of the NMB1880 divergent promoter, suggesting a role in its regulation. Mutants of N. meningitidis strains lacking MpeR or overexpressing MpeR showed no significant differences in expression of the P(NMB1880) promoter, nor did global transcriptional profiling of an MpeR knockout identify any deregulated genes, suggesting that the MpeR protein is inactive under the conditions used in these experiments. The presence of MpeR in a regulatory cascade downstream of the Fur master iron regulator implicates it as being expressed in the iron-limiting environment of the host, where it may in turn regulate a group of genes, including the divergent iron transport locus, in response to signals important for infection.
Staphylococcus aureus is a frequent cause for serious, chronic and therapy-refractive infections in spite of susceptibility to antibiotics in vitro. In chronic infections, altered bacterial phenotypes, such as small colony variants (SCVs), have been found. Yet, it is largely unclear whether the ability to interconvert from the wild-type to the SCV phenotype is only a rare clinical and/or just laboratory phenomenon or is essential to sustain an infection. Here, we performed different long-term in vitro and in vivo infection models with S. aureus and we show that viable bacteria can persist within host cells and/or tissues for several weeks. Persistence induced bacterial phenotypic diversity, including SCV phenotypes, accompanied by changes in virulence factor expression and auxotrophism. However, the recovered SCV phenotypes were highly dynamic and rapidly reverted to the fully virulent wild-type form when leaving the intracellular location and infecting new cells. Our findings demonstrate that bacterial phenotype switching is an integral part of the infection process that enables the bacteria to hide inside host cells, which can be a reservoir for chronic and therapy-refractive infections.
→ See accompanying Closeup by Christian Garzoni and William Kelley DOI 10.1002/emmm.201100123.
The mecA gene, encoding methicillin resistance in staphylococci, is located on a mobile genetic element called Staphylococcal Cassette Chromosome mec (SCCmec). Horizontal, interspecies transfer of this element could be an important factor in the dissemination of methicillin-resistant S. aureus (MRSA). Previously, we reported the isolation of a closely related methicillin-susceptible Staphylococcus aureus (MSSA), MRSA and potential SCCmec donor Staphylococcus epidermidis isolate from the same patient. Based on fingerprint techniques we hypothesized that the S. epidermidis had transferred SCCmec to the MSSA to become MRSA. The aim of this study was to show that these isolates form an isogenic pair and that interspecies horizontal SCCmec transfer occurred.
Whole genome sequencing of both isolates was performed and for the MSSA gaps were closed by conventional sequencing. The SCCmec of the S. epidermidis was also sequenced by conventional methods. The results show no difference in nucleotide sequence between the two isolates except for the presence of SCCmec in the MRSA. The SCCmec of the S. epidermidis and the MRSA are identical except for a single nucleotide in the ccrB gene, which results in a valine to alanine substitution. The main difference with the closely related EMRSA-16 is the presence of SaPI2 encoding toxic shock syndrome toxin and exfoliative toxin A in the MSSA-MRSA pair. No transfer of SCCmec from the S. epidermidis to the MSSA could be demonstrated in vitro.
The MSSA and MRSA form an isogenic pair except for SCCmec. This strongly supports our hypothesis that the MRSA was derived from the MSSA by interspecies horizontal transfer of SCCmec from S. epidermidis O7.1.
Staphylococcus aureus frequently invades the human bloodstream, leading to life threatening bacteremia and often secondary foci of infection. Failure of antibiotic therapy to eradicate infection is frequently described; in some cases associated with altered S. aureus antimicrobial resistance or the small colony variant (SCV) phenotype. Newer antimicrobials, such as linezolid, remain the last available therapy for some patients with multi-resistant S. aureus infections. Using comparative and functional genomics we investigated the molecular determinants of resistance and SCV formation in sequential S. aureus isolates from a patient who had a persistent and recurrent S. aureus infection, after failed therapy with multiple antimicrobials, including linezolid. Two point mutations in key staphylococcal genes dramatically affected clinical behaviour of the bacterium, altering virulence and antimicrobial resistance. Most strikingly, a single nucleotide substitution in relA (SACOL1689) reduced RelA hydrolase activity and caused accumulation of the intracellular signalling molecule guanosine 3', 5'-bis(diphosphate) (ppGpp) and permanent activation of the stringent response, which has not previously been reported in S. aureus. Using the clinical isolate and a defined mutant with an identical relA mutation, we demonstrate for the first time the impact of an active stringent response in S. aureus, which was associated with reduced growth, and attenuated virulence in the Galleria mellonella model. In addition, a mutation in rlmN (SACOL1230), encoding a ribosomal methyltransferase that methylates 23S rRNA at position A2503, caused a reduction in linezolid susceptibility. These results reinforce the exquisite adaptability of S. aureus and show how subtle molecular changes cause major alterations in bacterial behaviour, as well as highlighting potential weaknesses of current antibiotic treatment regimens.
MRSA, Close and Personal
Methods for differentiating pathogen isolates are essential for understanding their evolution and spread, as well as for the formulation of effective clinical strategies. Current typing methods for bacterial pathogens focus on a limited set of characteristics providing data with limited resolving power. Harris et al. (p. 469 ) used a high-throughput genome sequencing approach to show that isolates of methicillin-resistant Staphylococcus aureus (MRSA) are precisely differentiated into a global geographic structure. The findings suggest that intercontinental transmission has occurred for nearly four decades. The method could also detect individual person-to-person transmission events of MRSA within a hospital environment.
Although Staphylococcus aureus is a bacterial species of medical significance, only approximately 30% of all humans carry staphylococcal cells persistently but asymptomatically in their nasopharynx and/or other body sites. This goes largely unnoticed by the host, which shows that in the natural situation the human ecosystem is hospitable or at least receptive to the bacteria and that by a process of co-evolution this has lead to a state of mutual acceptance or tolerance. However, upon disturbance of this balanced, neutral state, localized or disseminated invasive infection can occur. Unfortunately, the events leading to infection are still largely unknown and especially the causal events leading to the transition from colonization to infection are ill-defined in vivo. Whether certain genotypes of S. aureus are more prone to colonise and/or infect humans is still quite heavily debated. The genetic population structure of S. aureus has been largely solved by using a number of different DNA polymorphism-interrogating laboratory methods. However, even this major effort has not (yet) revealed major clues with respect to colonisation and infection potency of the clonal lineages that were thus identified, except for the fact that certain lineages are highly epidemic. The overall picture is that in principle all S. aureus strains can become invasive given the proper circumstances. What these, primarily host-defined circumstances are is still enigmatic. However, a large variety of staphylococcal virulence and colonization factors have been identified as well as a number of host' colonisation and infection susceptibility traits. How these are specifically involved in colonisation and infection has been experimentally substantiated in only a limited number of cases. The present review paper will explore the relevance of these and other, for instance environmental factors that define the colonisation or infection state in humans. When the nature of these states would be known in more detail, this knowledge could be used to design novel and empirical, knowledge-driven means of preventing colonisation from proceeding into S. aureus infection.
The bacterial species Staphylococcus aureus, including its methicillin-resistant variant (MRSA), finds its primary ecological niche in the human nose, but is also able to colonize the intestines and the perineal region. Intestinal carriage has not been widely investigated despite its potential clinical impact. This review summarizes literature on the topic and sketches the current state of affairs from a microbiological and infectious diseases' perspective. Major findings are that the average reported detection rate of intestinal carriage in healthy individuals and patients is 20% for S. aureus and 9% for MRSA, which is approximately half of that for nasal carriage. Nasal carriage seems to predispose to intestinal carriage, but sole intestinal carriage occurs relatively frequently and is observed in 1 out of 3 intestinal carriers, which provides a rationale to include intestinal screening for surveillance or in outbreak settings. Colonization of the intestinal tract with S. aureus at a young age occurs at a high frequency and may affect the host's immune system. The frequency of intestinal carriage is generally underestimated and may significantly contribute to bacterial dissemination and subsequent risk of infections. Whether intestinal rather than nasal S. aureus carriage is a primary predictor for infections is still ill-defined.
The accessory gene regulator (agr) of Staphylococcus aureus is a global regulator of the staphylococcal virulon, which includes secreted virulence factors and surface proteins. The agr locus is important for virulence in a variety of animal models of infection, and has been assumed by inference to have a major role in human infection. Although most human clinical S. aureus isolates are agr(+), there have been several reports of agr-defective mutants isolated from infected patients. Since it is well known that the agr locus is genetically labile in vitro, we have addressed the question of whether the reported agr-defective mutants were involved in the infection or could have arisen during post-isolation handling. We obtained a series of new staphylococcal isolates from local clinical infections and handled these with special care to avoid post-isolation mutations. Among these isolates, we found a number of strains with non-haemolytic phenotypes owing to mutations in the agr locus, and others with mutations elsewhere. We have also obtained isolates in which the population was continuously heterogeneous with respect to agr functionality, with agr(+) and agr(-) variants having otherwise indistinguishable chromosomal backgrounds. This finding suggested that the agr(-) variants arose by mutation during the course of the infection. Our results indicate that while most clinical isolates are haemolytic and agr(+), non-haemolytic and agr(-) strains are found in S. aureus infections, and that agr(+) and agr(-) variants may have a cooperative interaction in certain types of infections.
A molecular epidemiological analysis was undertaken to identify lineages of Staphylococcus aureus that may be disproportionately associated with infection. Pulsed-field gel electrophoresis analysis of 405 S. aureus clinical isolates collected from various infection types and geographic locations was performed. Five distinct S. aureus lineages (SALs 1, 2, 4, 5, and 6) were identified, which accounted for 19.01, 9.14, 22.72, 10.12, and 4.69% of isolates,
respectively. In addition, 85 lineages which occurred with frequencies of <2.5% were identified and were termed “sporadic.”
The most prevalent lineage was methicillin-resistant S. aureus (SAL 4). The second most prevalent lineage, SAL 1, was also isolated at a high frequency from the anterior nares of healthy
volunteers, suggesting that its prevalence among clinical isolates may be a consequence of high carriage rates in humans.
Gene-specific PCR was carried out to detect genes for a number of staphylococcal virulence traits. tstand cna were found to be significantly associated with prevalent lineages compared to sporadic lineages. When specific infection
sites were examined, SAL 4 was significantly associated with respiratory tract infection, while SAL 2 was enriched among blood
isolates. SAL 1 and SAL 5 were clonally related to SALs shown by others to be widespread in the clinical isolate population.
We conclude from this study that at least five phylogenetic lineages of S. aureus are highly prevalent and widely distributed among clinical isolates. The traits that confer on these lineages a propensity
to infect may suggest novel approaches to antistaphylococcal therapy.
Vancomycin-resistant S. aureus was cultured from the exit site of a temporary dialysis catheter of a patient with diabetes, peripheral vascular disease, and persistent foot ulcers. The organism carried the vanA resistance gene, which may have come from the vancomycin-resistant Enterococcus faecalis that was also isolated from the patient, who had received vancomycin for several weeks.
Staphylococcus aureus is an important human pathogen and represents a growing public health burden owing to the emergence and spread of antibiotic-resistant clones, particularly within the hospital environment. Despite this, basic questions about the evolution and population biology of the species, particularly with regard to the extent and impact of homologous recombination, remain unanswered. We address these issues through an analysis of sequence data obtained from the characterization by multilocus sequence typing (MLST) of 334 isolates of S. aureus, recovered from a well-defined population, over a limited time span. We find no significant differences in the distribution of multilocus genotypes between strains isolated from carriers and those from patients with invasive disease; there is, therefore, no evidence from MLST data, which index variation within the stable "core" genome, for the existence of hypervirulent clones of this pathogen. Examination of the sequence changes at MLST loci during clonal diversification shows that point mutations give rise to new alleles at least 15-fold more frequently than does recombination. This contrasts with the naturally transformable species Neisseria meningitidis and Streptococcus pneumoniae, in which alleles change between 5- and 10-fold more frequently by recombination than by mutation. However, phylogenetic analysis suggests that homologous recombination does contribute toward the evolution of this species over the long term. Finally, we note a striking excess of nonsynonymous substitutions in comparisons between isolates belonging to the same clonal complex compared to isolates belonging to different clonal complexes, suggesting that the removal of deleterious mutations by purifying selection may be relatively slow.
Vancomycin is usually reserved for treatment of serious infections, including those caused by multidrug-resistant Staphylococcus aureus. A clinical isolate of S. aureus with high-level resistance to vancomycin (minimal inhibitory concentration = 1024 microg/ml) was isolated in June 2002. This isolate harbored a 57.9-kilobase multiresistance conjugative plasmid within which Tn1546 (vanA) was integrated. Additional elements on the plasmid encoded resistance to trimethoprim (dfrA), beta-lactams (blaZ), aminoglycosides (aacA-aphD), and disinfectants (qacC). Genetic analyses suggest that the long-anticipated transfer of vancomycin resistance to a methicillin-resistant S. aureus occurred in vivo by interspecies transfer of Tn1546 from a co-isolate of Enterococcus faecalis.
We assessed the nature and frequency of genome alterations in Staphylococcus aureus during chronic lung infection in patients with cystic fibrosis (CF) and during colonization of the nares in healthy individuals.
Only individuals harboring the same S. aureus clone on consecutive samplings were included in the present study. Clone definition was based on pulsed-field gel electrophoresis
(PFGE) analysis. Minor fragment variations in consecutive clones were interpreted as genome alterations. The frequency of
genome alterations was significantly higher in S. aureus derived from patients with CF (mean time, 1.03 years) than in isolates derived from healthy individuals (mean time, 13.4
years). In total, 19 S. aureus strain pairs showing genome alterations were available for molecular analysis to clarify the nature of recombinational events
in the host environment. In 8 cases, genome alteration could be linked to phage mobilization. Phage conversion of β-toxin
production was evident in 7 pairs. In 1 strain pair, changes in the PFGE pattern were accompanied by deletion of a phage similar
to ETA. Obviously, phage mobilization plays an important role in vivo. During long-termlung infection in patients with CF,
the specific host response and/or the regular exposure to antibiotics exercises strong selective pressure on the pathogen.
Genome plasticity may facilitate the adaptation to various host conditions.
Variation of the polymorphic region of the protein A gene (spa) was observed during long-term persistence of Staphylococcus aureus in the airways of 10 cystic fibrosis patients and occurred at a rate of one genetic change every 70 months. Independent mutational
events were observed eight times in 142 isolates: four deletions, two duplications of repeats, and two point mutations.
Staphylococcus aureus is a frequent cause of infections in both the community and hospital. Worldwide, the increasing resistance of this pathogen to various antibiotics complicates treatment of S aureus infections. Effective measures to prevent S aureus infections are therefore urgently needed. It has been shown that nasal carriers of S aureus have an increased risk of acquiring an infection with this pathogen. The nose is the main ecological niche where S aureus resides in human beings, but the determinants of the carrier state are incompletely understood. Eradication of S aureus from nasal carriers prevents infection in specific patient categories-eg, haemodialysis and general surgery patients. However, recent randomised clinical trials in orthopaedic and non-surgical patients failed to show the efficacy of eliminating S aureus from the nose to prevent subsequent infection. Thus we must elucidate the mechanisms behind S aureus nasal carriage and infection to be able to develop new preventive strategies. We present an overview of the current knowledge of the determinants (both human and bacterial) and risks of S aureus nasal carriage. Studies on the population dynamics of S aureus are also summarised.
Staphylococcus aureus is the most common cause of hospital-acquired infection. In healthy hosts outside of the health care setting, S. aureus is a frequent colonizer of the human nose but rarely causes severe invasive infection such as bacteremia, endocarditis, or
osteomyelitis. To identify genes associated with community-acquired invasive isolates, regions of genomic variability, and
the S. aureus population structure, we compared 61 community-acquired invasive isolates of S. aureus and 100 nasal carriage isolates from healthy donors using a microarray spotted with PCR products representing every gene
from the seven S. aureus sequencing projects. The core genes common to all strains were identified, and 10 dominant lineages of S. aureus were clearly discriminated. Each lineage carried a unique combination of hundreds of “core variable” (CV) genes scattered
throughout the chromosome, suggesting a common ancestor but early evolutionary divergence. Many CV genes are regulators of
virulence genes or known or predicted to be expressed on the bacterial surface and to interact with the host during nasal
colonization and infection. Within each lineage, isolates showed substantial variation in the carriage of mobile genetic elements
and their associated virulence and resistance genes, indicating frequent horizontal transfer. However, we were unable to identify
any association between lineage or gene and invasive isolates. We suggest that the S. aureus gene combinations necessary for invasive disease may also be necessary for nasal colonization and that community-acquired
invasive disease is strongly dependent on host factors.
Small colony variants constitute a slow-growing subpopulation of bacteria with distinctive phenotypic and pathogenic traits. Phenotypically, small colony variants have a slow growth rate, atypical colony morphology and unusual biochemical characteristics, making them a challenge for clinical microbiologists to identify. Clinically, small colony variants are better able to persist in mammalian cells and are less susceptible to antibiotics than their wild-type counterparts, and can cause latent or recurrent infections on emergence from the protective environment of the host cell. This Review covers the phenotypic, genetic and clinical picture associated with small colony variants, with an emphasis on staphylococci, for which the greatest amount of information is available.
Daptomycin is a lipopeptide antibiotic with potent activity against gram-positive bacteria. Complete-genome comparisons of
laboratory-derived Staphylococcus aureus with decreased susceptibility to daptomycin and their susceptible parent were used to identify genes that contribute to reduced
susceptibility to daptomycin. Selective pressure of growth in sublethal concentrations of daptomycin resulted in the accumulation
of mutations over time correlating with incremental decreases in susceptibility. Single point mutations resulting in amino
acid substitutions occurred in three distinct proteins: MprF, a lysylphosphatidylglycerol synthetase; YycG, a histidine kinase;
and RpoB and RpoC, the β and β′ subunits of RNA polymerase. Sequence analysis of mprF, yycF, yycG, rpoB, and rpoC in clinical isolates that showed treatment-emergent increases in daptomycin MICs revealed point mutations in mprF and a nucleotide insertion in yycG, suggesting a role for these genes in decreased susceptibility to daptomycin in the hospital setting.
The Sau1 type I restriction-modification system is found on the chromosome of all nine sequenced strains of Staphylococcus aureus and includes a single hsdR (restriction) gene and two copies of hsdM (modification) and hsdS (sequence specificity) genes. The strain S. aureus RN4220 is a vital intermediate for laboratory S. aureus manipulation, as it can accept plasmid DNA from Escherichia coli. We show that it carries a mutation in the sau1hsdR gene and that complementation restored a nontransformable phenotype. Sau1 was also responsible for reduced conjugative transfer
from enterococci, a model of vancomycin resistance transfer. This may explain why only four vancomycin-resistant S. aureus strains have been identified despite substantial selective pressure in the clinical setting. Using a multistrain S. aureus microarray, we show that the two copies of sequence specificity genes (sau1hsdS1 and sau1hsdS2) vary substantially between isolates and that the variation corresponds to the 10 dominant S. aureus lineages. Thus, RN4220 complemented with sau1hsdR was resistant to bacteriophage lysis but only if the phage was grown on S. aureus of a different lineage. Similarly, it could be transduced with DNA from its own lineage but not with the phage grown on different
S. aureus lineages. Therefore, we propose that Sau1 is the major mechanism for blocking transfer of resistance genes and other mobile
genetic elements into S. aureus isolates from other species, as well as for controlling the spread of resistance genes between isolates of different S. aureus lineages. Blocking Sau1 should also allow genetic manipulation of clinical strains of S. aureus.
Horizontal genetic transfer (HGT) has played an important role in bacterial evolution at least since the origins of the bacterial divisions, and HGT still facilitates the origins of bacterial diversity, including diversity based on antibiotic resistance. Adaptive HGT is aided by unique features of genetic exchange in bacteria such as the promiscuity of genetic exchange and the shortness of segments transferred. Genetic exchange rates are limited by the genetic and ecological similarity of organisms. Adaptive transfer of genes is limited to those that can be transferred as a functional unit, provide a niche-transcending adaptation, and are compatible with the architecture and physiology of other organisms. Horizontally transferred adaptations may bring about fitness costs, and natural selection may ameliorate these costs. The origins of ecological diversity can be analyzed by comparing the genomes of recently divergent, ecologically distinct populations, which can be discovered as sequence clusters. Such genome comparisons demonstrate the importance of HGT in ecological diversification. Newly divergent populations cannot be discovered as sequence clusters when their ecological differences are coded by plasmids, as is often the case for antibiotic resistance; the discovery of such populations requires a screen for plasmid-coded functions. This paper reviews the features of bacterial genetics that allow HGT, the similarities between organisms that foster HGT between them, the limits to the kinds of adaptations that can be transferred, and amelioration of fitness costs associated with HGT; the paper also reviews approaches to discover the origins of new, ecologically distinct bacterial populations and the role that HGT plays in their founding.
This review highlights the exciting new finding that small molecule inducers have been found for some members of the AraC family of transcriptional regulators (AFTRs) that control the expression of virulence genes of pathogenic bacteria. Although effector-mediated activation of AFTRs involved in the regulation of sugar and alkylbenzoate catabolism (e.g. AraC and XylS) is well understood, until recently little was known about effector-mediated regulation of virulence gene expression. This led to the belief that regulation of virulence by AFTRs does not involve the direct recognition of chemical environmental signals. More recently, however, a growing number of virulence-associated AFTRs have been found to directly sense environmental chemicals. Most interestingly, these environmental chemicals are abundant at the sites where the bacterial pathogen colonizes and damages its host. In this article we review recent developments in the field of environmental regulation mediated by virulence-associated AFTRs, with a focus on the sensing of environmental signals, the mechanism of gene target activation, and the effector-mediated modulation of regulator activities.
Staphylococcus aureus is thought to have acquired mecA DNA by horizontal transfer. DNA fingerprints made by restriction nucleases that cut certain sequences of DNA are used to compare complete genomes or particular genes between bacteria. We isolated an epidemic mecA− meticillin-susceptible S aureus genotype and, subsequently, a rare isogeneic mecA+ meticillin-resistant S aureus (MRSA) genotype from a neonate who had never been in contact with MRSA. This MRSA contained mecA DNA that was identical to that in a coagulase-negative staphylococcal strain isolated from this patient, but different from other MRSA genotypes. We believe that this MRSA was formed in vivo by horizontal transfer of the mecA DNA between two staphylococcal species.
One of the first pathogens which can be isolated from the airways of cystic fibrosis (CF) patients is Staphylococcus aureus, which often persists in this hostile environment for many months or even years. The increase in infections due to the methicillin-resistant S. aureus (MRSA) worldwide and even more the emergence of community-acquired MRSA, which differ from nosocomial MRSA by lack of multiresistance and carriage of a phage-encoded toxin, Panton-Valentine leukocidin, attracts new attention to the epidemiology, pathogenesis, and impact of S. aureus in the background of CF. In this review, recent data and studies will be reported and discussed to give an overview of the latest research.
Staphylococcus aureus is an important human pathogen of endovascular diseases, which can take a chronic course with a high relapse rate despite antimicrobial treatment. Thus far, persistent and antibiotic-refractory infections have been largely associated with a subpopulation of S. aureus, the small-colony variants (SCVs).
In this work, we used endothelial cells to investigate infection with the highly virulent wild-type isolate (6850), 2 stable isogenic SCV phenotypes (hemB mutant IIb13 and JB1), and the complemented mutant.
All strains were highly invasive in endothelial cells but largely differed in host response induction. Microarray analysis showed that wild-type phenotypes up-regulated a large number of endothelial genes (including genes involved in innate immunity), whereas the SCVs did not cause these dramatic changes. The inflammatory response and cytotoxicity were strongest shortly after infection and largely decreased within the following days, which was accompanied by a fast elimination of intracellular wild-type bacteria. By contrast, SCVs survived within endothelial cells at high numbers.
S. aureus intracellular persistence via the development of an adapted subpopulation of SCVs most likely represents an important strategy of S. aureus to hide within the host cells, which could be a reservoir for chronic infections.
Staphylococcus aureus is the leading cause of bacterial infections in developed countries and produces a wide spectrum of diseases, ranging from minor skin infections to fatal necrotizing pneumonia. Although S. aureus infections were historically treatable with common antibiotics, emergence of drug-resistant organisms is now a major concern. Methicillin-resistant S. aureus (MRSA) was endemic in hospitals by the late 1960s, but it appeared rapidly and unexpectedly in communities in the 1990s and is now prevalent worldwide. This Review focuses on progress made toward understanding the success of community-associated MRSA as a human pathogen, with an emphasis on genome-wide approaches and virulence determinants.
Staphylococcus aureus is notorious for its ability to become resistant to antibiotics. Infections that are caused by antibiotic-resistant strains often occur in epidemic waves that are initiated by one or a few successful clones. Methicillin-resistant S. aureus (MRSA) features prominently in these epidemics. Historically associated with hospitals and other health care settings, MRSA has now emerged as a widespread cause of community infections. Community or community-associated MRSA (CA-MRSA) can spread rapidly among healthy individuals. Outbreaks of CA-MRSA infections have been reported worldwide, and CA-MRSA strains are now epidemic in the United States. Here, we review the molecular epidemiology of the epidemic waves of penicillin- and methicillin-resistant strains of S. aureus that have occurred since 1940, with a focus on the clinical and molecular epidemiology of CA-MRSA.
Strains of Staphylococcus aureus that are resistant to multiple antimicrobial compounds, including most available classes of antibiotics and some antiseptics, are a major threat to patient care owing to their stubborn intransigence to chemotherapy and disinfection. This reality has stimulated extensive efforts to understand the genetic nature of the determinants encoding antimicrobial resistance, together with the mechanisms by which these determinants evolve over time and are spread within bacterial populations. Such studies have benefited from the application of molecular genetics and in recent years, the sequencing of over a dozen complete staphylococcal genomes. It is now evident that the evolution of multiresistance is driven by the acquisition of discrete preformed antimicrobial resistance genes that are exchanged between organisms via horizontal gene transfer. Nonetheless, chromosomal mutation is the catalyst of novel resistance determinants and is likely to have an enhanced influence with the ongoing introduction of synthetic antibiotics.
Persistent nasal carriers have an increased risk of Staphylococcus aureus infection, whereas intermittent carriers and noncarriers share the same low risk. This study was performed to provide additional insight into staphylococcal carriage types.
Fifty-one volunteers who had been decolonized with mupirocin treatment and whose carriage state was known were colonized artificially with a mixture of S. aureus strains, and intranasal survival of S. aureus was compared between carriage groups. Antistaphylococcal antibody levels were also compared among 83 carriage-classified volunteers.
Persistent carriers preferentially reselected their autologous strain from the inoculum mixture (P=.02). They could be distinguished from intermittent carriers and noncarriers on the basis of the duration of postinoculation carriage (154 vs. 14 and 4 days, respectively; P=.017, by log-rank test). Cultures of swab samples from persistent carriers contained significantly more colony-forming units per sample than did cultures of swab samples from intermittent carriers and noncarriers (P=.004). Analysis of serum samples showed that levels of immunoglobulin G and immunoglobulin A to 17 S. aureus antigens were equal in intermittent carriers and noncarriers but not in persistent carriers.
Along with the previously described low risk of infection, intermittent carriers and noncarriers share similar S. aureus nasal elimination kinetics and antistaphylococcal antibody profiles. This implies a paradigm shift; apparently, there are only 2 types of nasal carriers: persistent carriers and others. This knowledge may increase our understanding of susceptibility to S. aureus infection.
Because of its importance as a major human and animal pathogen, Staphylococcus aureus has been the focus of intensive research efforts. At the time of writing, the genomes of 14 isolates of S. aureus have been published and released into the public domain with many more genome sequencing projects underway, representing an excellent resource for studies of bacterial evolution and pathogenesis. Recently, whole genome sequences of several other species of the Staphylococcus genus have been completed allowing a comparative genomic analysis of the adaptation of different species to their natural habitats. Here, we summarize selected comparative genomic studies that have contributed to our understanding of how staphylococci adapt to different environments, combat antibiotics and acquire increased virulence.
The first methicillin-resistant Staphylococcus aureus (MRSA) strain originated when a staphylococcal cassette chromosome mec (SCCmec) with the gene mecA was integrated into the chromosome of a susceptible S. aureus cell. The SCCmec elements are common among the coagulase-negative staphylococci, e.g. Staphylococcus haemolyticus, and these are considered to be potential SCCmec donors when new clones of MRSA arise. An outbreak of MRSA occurred at a neonatal intensive-care unit, and the isolates were all of sequence type (ST) 45, as characterized by multilocus sequence typing, but were not typeable with respect to SCCmec types I, II, III or IV. During the same time period, methicillin-resistant S. haemolyticus (MRSH) isolates identified in blood cultures at the same ward were found to be genotypically homogenous by pulsed-field gel electrophoresis, and did not carry a type I, II, III or IV SCCmec either. Thus, the hypothesis was raised that an SCCmec of MRSH had been transferred to a methicillin-susceptible S. aureus strain and thereby created a new clone of MRSA that caused the outbreak. This study showed that MRSA from the outbreak carried a ccrC and a class C mec complex that was also found among MRSH isolates. Partial sequencing of the mec complexes showed more than 99% homology, indicative of a common type V SCCmec. This finding may provide evidence for a recent horizontal transfer of an SCCmec from MRSH to an identified potential recipient, an ST45 methicillin-susceptible S. aureus strain, thereby creating a new clone of MRSA that caused the outbreak.
Microbial pathogenicity in Staphylococcus aureus is a complex process involving a number of virulence genes that are regulated by global regulatory systems including sar and agr. To evaluate the roles of these two loci in virulence, we constructed sar-/agr- mutants of strains RN6390 and RN450 and compared their phenotypic profiles to the corresponding single sar- and agr- mutants and parents. The secretion of all hemolysins was absent in the sar-/agr- mutants while residual beta-hemolysin activity remained in single agr- mutants. The fibronectin binding capacity was significantly diminished in both single sar- mutants and double mutants when compared with parents while the reduction in fibrinogen binding capacity in the double mutants was modest. In the rabbit endocarditis model, there was a significant decrease in both infectivity rates and intravegetation bacterial densities with the double mutant as compared to the parent (RN6390) at 10(3)-10(6) CFU inocula despite comparable levels of early bacteremia among various challenge groups. Notably, fewer bacteria in the double mutant group adhered to valvular vegetations at 30 min after challenge (10(6) CFU) than the parent group. These studies suggest that both the sar and agr loci are involved in initial valvular adherence, intravegetation persistence and multiplication of S. aureus in endocarditis.
We have studied the role of the accessory gene regulator (agr) of Staphylococcus aureus as a virulence determinant in the pathogenesis of septic arthritis. At least 15 genes coding for potential virulence factors in Staphylococcus aureus are regulated by a putative multicomponent signal transduction system encoded by the agr/hld locus. agr and hld mutants show a decreased synthesis of extracellular toxins and enzymes, such as alpha-, beta-, and delta-hemolysin, leucocidin, lipase, hyaluronate lyase, and proteases, and at the same time an increased synthesis of coagulase and protein A as compared with the wild-type counterpart. We have used a recently described murine model of S. aureus-induced arthritis to study the virulence of S. aureus 8325-4 and two agr/hld mutants derived from it. Sixty percent of the mice injected with the wild-type strain developed arthritis, whereas agrA and hld mutants displayed joint involvement in only 10 and 30%, respectively. In addition, 40% of the mice inoculated with the wild-type strain displayed an erosive arthropathy; such changes were not detectable at all in mice inoculated with the agrA mutant. Serum levels of interleukin-6, a potent B-cell differentiation factor, were significantly higher (P < 0.001) in the mice inoculated with the wild-type strain than in those inoculated with the agrA mutant counterpart. Overall, our results suggest that the agr system of S. aureus is an important virulence determinant in the induction and progression of septic arthritis in mice.
Pulsed-field gel electrophoresis (PFGE) of SmaI macrorestriction fragments of chromosomal DNA was used to confirm the persistence of methicillin-sensitive Staphylococcus aureus isolates in the sputum of 25 cystic fibrosis patients in five French hospitals. Three-to-eight consecutive isolates, with the same esterase electrophoretic type isolated from each patient over a period of 12-28 months, were analysed. Consecutive isolates with indistinguishable PFGE profiles were found in 12 patients (48%) and consecutive isolates with similar PFGE profiles showing minor differences of one-to-four fragments (similarity coefficient >/=84%) were found in 11 patients. Consecutive isolates with different PFGE profiles were obtained from only two patients, but the profiles found in each patient were more closely related to each other than to other profiles. The results were in agreement with esterase electrophoretic typing for 23 patients, and we considered that those patients were infected with a single persistent strain. For any given patient, variations in antibiotypes and phage types of consecutive isolates were not associated with major genotypic variations. PFGE is useful in confirming the persistence of S. aureus strains in cystic fibrosis patients over long periods.
Micrococcus, which, when limited in its extent and activity, causes acute suppurative inflammation (phlegmon), produces, when more extensive and intense in its action on the human system, the most virulent forms of septicaemia and pyaemia.1 In an elegant series of clinical observations and laboratory studies published in 1880 and 1882, Ogston described staphylococcal disease and its role in sepsis and abscess formation.1,2 More than 100 years later, Staphylococcus aureus remains a versatile and dangerous pathogen in humans. The frequencies of both community-acquired and hospital-acquired staphylococcal infections have increased steadily, with little change in overall mortality. Treatment of these infections . . .
We previously reported the first vancomycin-resistant Staphylococcus aureus (VRSA) clinical strain, Mu50, whose cell wall is remarkably thickened resulting from the activation of cell-wall synthesis. To explore the genetic basis for the vancomycin resistance, cDNA differential hybridization was performed using RNAs extracted from a set of closely related S. aureus strains with various levels of vancomycin susceptibilities. The strains were Mu3 (MIC = 2 microg/ml), Mu50 (MIC = 8 microg/ml), and a susceptible revertant of Mu50, Mu50omega (MIC = 0.5 microg/ml). In this study, we report identification of a novel response regulator, designated vraR (standing for vancomycin-resistance associated gene R) whose transcription was remarkably up-regulated in Mu3 and Mu50 as compared to Mu50omega. Experimental over-expression of VraR in vancomycin-susceptible strain N315P raised vancomycin resistance of the strain. Also, the genes coding for fructose utilization, fatty acid metabolism, and two putative ATP-binding cassette (ABC) transporter genes were found to be up-regulated in Mu3 and Mu50. On the other hand, Protein A expression was suppressed in Mu50, as compared with Mu3 and Mu50omega. We consider that the response regulator vraR is one of the key regulators modulating the level of vancomycin-resistance in S. aureus. Presumed increased uptake of fructose and altered fatty acid metabolism may also contribute to vancomycin resistance by supplying more precursor metabolites for cell-wall synthesis.
Staphylococcus aureus is thought to have acquired mecA DNA by horizontal transfer. DNA fingerprints made by restriction nucleases that cut certain sequences of DNA are used to compare complete genomes or particular genes between bacteria. We isolated an epidemic mecA(-) meticillin-susceptible S aureus genotype and, subsequently, a rare isogeneic mecA(+) meticillin-resistant S aureus (MRSA) genotype from a neonate who had never been in contact with MRSA. This MRSA contained mecA DNA that was identical to that in a coagulase-negative staphylococcal strain isolated from this patient, but different from other MRSA genotypes. We believe that this MRSA was formed in vivo by horizontal transfer of the mecA DNA between two staphylococcal species.
The causes of persistent bacteremia (PB) due to methicillin-resistant Staphylococcus aureus (MRSA) are poorly understood. This investigation examined potential associations between PB with key clinical features and several in vitro bacterial genotypic and phenotypic characteristics, in isolates from 1 institution.
Pulsed-field gel electrophoresis (PFGE) relatedness, thrombin-induced platelet microbicidal protein (tPMP)-susceptibility phenotype, accessory gene regulator (agr) genotype and functionality (via delta-lysin production), and autolysis phenotypes were assessed in MRSA isolates from the bloodstream of 21 prospectively identified patients with PB (blood cultures positive after > or =7 days of therapy) and of 18 patients with resolving bacteremia (RB) (sterile blood cultures within the first 2-4 days of therapy) due to MRSA.
The 2 groups had comparable baseline characteristics but differed in their clinical courses (e.g., endocarditis was more frequent in patients with PB than in those with RB [43% vs. 0%, respectively; P=.0016]); isolates from patients with PB exhibited higher rates of (1) survival in vitro after exposure to tPMP (22.4+/-14.8% vs. 11.6+/-6.5%, respectively; P=.005); (2) defective delta-lysin production (71.4% vs. 38.9%, respectively; P=.057); (3) non-agr genotype II profile (100% vs. 77.8%, respectively; P=.037); and (4) overrepresentation of a specific PFGE genotype (85.7% vs. 44.4%, respectively; P=.015).
Isolates from patients with PB differed from those in patients with RB, in several in vitro characteristics. Further studies will be necessary to define how these factors might affect clinical outcome.
Staphylococcus aureus can cause superficial skin infections and, occasionally, deep-seated infections that entail spread through the blood stream. The organism expresses several factors that compromise the effectiveness of neutrophils and macrophages, the first line of defence against infection. S. aureus secretes proteins that inhibit complement activation and neutrophil chemotaxis or that lyse neutrophils, neutralizes antimicrobial defensin peptides, and its cell surface is modified to reduce their effectiveness. The organism can survive in phagosomes, express polysaccharides and proteins that inhibit opsonization by antibody and complement, and its cell wall is resistant to lysozyme. Furthermore, S. aureus expresses several types of superantigen that corrupt the normal humoral immune response, resulting in anergy and immunosuppression. In contrast, Staphylococcus epidermidis must rely primarily on cell-surface polymers and the ability to form a biolfilm to survive in the host.
Methicillin-resistant Staphylococcus aureus (MRSA) is well-recognised as a major cause of infection in the health care setting but, even more worryingly, is now emerging in the community. The glycopeptides-notably vancomycin-have traditionally been the mainstay of treatment of MRSA but overuse has led to the emergence of vancomycin-intermediate and vancomycin-resistant MRSA (VISA and VRSA, respectively). Although the mechanisms underlying vancomycin resistance are not yet fully understood, changes to the bacterial cell wall-the site of action of the glycopeptides-are believed to be key. Recent evidence also supports the transfer of genetic material among bacteria as contributing to the development of VRSA. Based on the cases identified to date, risk factors for the development of VRSA may include older age, compromised blood flow to the lower limbs, and the presence of chronic ulcers. The true extent of the problem, however, remains to be determined-it is likely that many cases of VISA and VRSA infection go undetected because of suboptimal screening programmes and possible limitations of automated and non-automated detection methods. Effective screening directed at those patients considered to be most at risk should therefore be a priority. Not surprisingly, the spread of MRSA from the hospital to the community setting, coupled with the emergence of VISA and VRSA, has become a major cause of concern among clinicians and microbiologists. The treatment options available for these infections are now severely compromised and thus new classes of antimicrobial agents effective against MRSA, VISA and VRSA are urgently required.