Staphylococcal Small Colony Variants Research Articles

Unravelling staphylococcal small-colony variants in cardiac implantable electronic device infections: clinical characteristics, management, and genomic insights.

Staphylococcal small-colony variants (SCVs) are common in cardiac implantable electronic device (CIED) infections. This is the first retrospective and multi-case study on CIED infections due to staphylococcal SCVs, aiming to provide a theoretical basis for the clinical management of CIED and device-related infections caused by staphylococcal SCVs. Ninety patients with culture positive CIED infections were enrolled between 2021 and 2022. We compared the demographic and clinical characteristics of patients with and without SCVs and performed genomic studies on SCVs isolates. Compared to patients without SCVs, those with SCVs had a longer primary pacemaker implantation time and were more likely to have a history of device replacement and infection. They showed upregulated inflammatory indicators, especially higher NEUT% (52.6 vs. 26.8%, P = 0.032) and they had longer hospital stays (median 13 vs. 12 days, P = 0.012). Comparative genomics analysis was performed on Staphylococcus epidermidis wild-type and SCVs. Some genes were identified, including aap, genes encoding adhesin, CHAP domain-containing protein, LPXTG cell wall anchor domain-containing protein, and YSIRK-type signal peptide-containing protein. Staphylococcal SCVs affect the clinical characteristics of CIED infections. The process of staphylococcal SCVs adherence, biofilm formation, and interaction with neutrophils play a vital role.

Synchrotron-Radiation-Based Fourier Transform Infrared Microspectroscopy as a Tool for the Differentiation between Staphylococcal Small Colony Variants.

Small colony variants (SCVs) are clinically significant and linked to persistent infections. In this study, synchrotron-radiation-based Fourier transform infrared (SR-FTIR) is used to investigate the microspectroscopic differences between the SCVs of Staphylococcus aureus (S. aureus) and diabetic foot Staphylococcus epidermidis (S. epidermidis) in two main IR spectral regions: (3050-2800 cm-1), corresponding to the distribution of lipids, and (1855-1500 cm-1), corresponding to the distribution of protein amide I and amide II and carbonyl vibrations. SR-FTIR successfully discriminated between the two staphylococcal species and between the SCV and the non-SCV strains within the two IR spectral regions. Combined S. aureus SCVs (SCVhMu) showed a higher protein content relative to the non-SCV wild type. Complemented S. aureus SCV showed distinguishable differences from the SCVhMu and the wild type, including a higher content of unsaturated fatty acids. An increase in the CH2/CH3 ratio was detected in S. epidermidis SCV samples compared to the standard control. Protein secondary structure in standard S. epidermidis and SCVs consisted mainly of an α-helix; however, a new shoulder at 1635 cm-1, assigned to β-sheets, was evident in the SCV. In conclusion, SR-FTIR is a powerful method that can discriminate between staphylococci species and to differentiate between SCVs and their corresponding natural strains.

MicroRNA hsa-let-7a facilitates staphylococcal small colony variants survival in the THP-1 macrophages by reshaping inflammatory responses

Recent studies have provided emerging evidence of the critical involvement of microRNAs in host immune defence against bacterial infection and that likewise the expression of the miRNAs is profoundly impacted by a variety of pathogens to subvert the immune response. Here, we report the role of hsa-let-7a miRNA in response to Staphylococcus epidermidis Small Colony Variants infection. We also assessed whether the expression levels of inflammatory cytokines associated with the hsa-let-7a are manipulated by the pathogen and the effect of the IFN-γ priming on the expression of hsa-let-7a and the fate of SCVs/WTs in infected macrophages. A striking observation was the downregulation of the let-7a miRNA upon challenge of the THP-1 activated cells with the SCV isolates while no significant changes in expression were noticed after the infection of macrophages with their WT counterparts. Staphylococcus epidermidis WT and SCV strains were found to invade and survive in macrophages. A significant reduction in bacterial load for both phenotypes was observed in macrophages treated with let-7a mimic compared to untreated ones. Survival of WTs was augmented in cells treated with the inhibitor in 4 out of 5 strains as compared to the number of bacteria recovered from non-transfected cells. At the same time, let-7a inhibitor did not influence on the survival of SCVs in macrophages as their number was comparable to number recovered from non-transfected cells. When the ratio of both let-7a cytokine targets was compared, anti-inflammatory IL-10 cytokine was induced by SCVs predominantly, while the macrophage challenge with WTs was characterized by the inflammatory cytokine profile with high IL-6 and low IL-10 production. Moreover, the balance between pro-inflammatory and anti-inflammatory cytokines has been expectedly retrieved when macrophages were transfected with let-7a mimic before infection with WT or SCV strains. The results also show that IFN-γ likely regulates the macrophage environment contributing to the inflammatory response and elimination of bacteria from intracellular milieu by augmenting the synthesis of pro-inflammatory cytokines and supressing the anti-inflammatory IL-10. Our work has shown that SCVs have the potential to regulate the let-7a miRNA to balance the pro-inflammatory IL-6 with anti-inflammatory IL-10 and this mechanism is one of the ways in a complex regulatory network adopted by SCVs to promote their survival.

The Energy-Coupling Factor Transporter Module EcfAA'T, a Novel Candidate for the Genetic Basis of Fatty Acid-Auxotrophic Small-Colony Variants of Staphylococcus aureus.

Staphylococcal small-colony variants (SCVs) are invasive and persistent due to their ability to thrive intracellularly and to evade the host immune response. Thus, the course of infections due to this phenotype is often chronic, relapsing, and therapy-refractory. In order to improve treatment of patients suffering from SCV-associated infections, it is of major interest to understand triggers for the development of this phenotype, in particular for strains naturally occurring in clinical settings. Within this study, we comprehensively characterized two different Staphylococcus aureus triplets each consisting of isogenic strains comprising (i) clinically derived SCV phenotypes with auxotrophy for unsaturated fatty acids, (ii) the corresponding wild-types (WTs), and (iii) spontaneous in vitro revertants displaying the normal phenotype (REVs). Comparison of whole genomes revealed that clinical SCV isolates were closely related to their corresponding WTs and REVs showing only seven to eight alterations per genome triplet. However, both SCVs carried a mutation within the energy-coupling factor (ECF) transporter-encoding ecf module (EcfAA’T) resulting in truncated genes. In both cases, these mutations were shown to be naturally restored in the respective REVs. Since ECF transporters are supposed to be essential for optimal bacterial growth, their dysfunction might constitute another mechanism for the formation of naturally occurring SCVs. Another three triplets analyzed revealed neither mutations in the EcfAA’T nor in other FASII-related genes underlining the high diversity of mechanisms leading to the fatty acid-dependent phenotype. This is the first report on the ECF transporter as genetic basis of fatty acid–auxotrophic staphylococcal SCVs.

Clinical Significance and Pathogenesis of Staphylococcal Small Colony Variants in Persistent Infections.

Small colony variants (SCVs) were first described more than 100 years ago for Staphylococcus aureus and various coagulase-negative staphylococci. Two decades ago, an association between chronic staphylococcal infections and the presence of SCVs was observed. Since then, many clinical studies and observations have been published which tie recurrent, persistent staphylococcal infections, including device-associated infections, bone and tissue infections, and airway infections of cystic fibrosis patients, to this special phenotype. By their intracellular lifestyle, SCVs exhibit so-called phenotypic (or functional) resistance beyond the classical resistance mechanisms, and they can often be retrieved from therapy-refractory courses of infection. In this review, the various clinical infections where SCVs can be expected and isolated, diagnostic procedures for optimized species confirmation, and the pathogenesis of SCVs, including defined underlying molecular mechanisms and the phenotype switch phenomenon, are presented. Moreover, relevant animal models and suggested treatment regimens, as well as the requirements for future research areas, are highlighted.

Staphylococcal small colony variants from a libyan hospital.

Staphylococcal small colony variants from a libyan hospital.

Intestinal staphylococcal small colony variants: a cause of medically unexplained physical symptoms?

No abstract available. (Published: 19 September 2014) Citation: Microbial Ecology in Health & Disease 2014, 25 : 25817 - http://dx.doi.org/10.3402/mehd.v25.25817

Characterization of Staphylococcus epidermidis and Staphyloccocus warneri small-colony variants associated with prosthetic-joint infections

We determined the frequency of isolation of staphylococcal small-colony variants (SCVs) from 31 culture-positive patients undergoing revision of total hip prosthesis for aseptic loosening or presumed prosthetic-joint infection (PJI). We analysed auxotrophy of cultured SCVs, their antimicrobial susceptibility profiles and their biofilm-forming capacity. Eight SCV strains were cultivated from six (19 %) patients. All SCVs were coagulase-negative staphylococci (CNS) with Staphylococcus epidermidis as the predominant species; there was also one Staphylococcus warneri SCV. The SCVs were auxotrophic for haemin, with one strain additionally auxotrophic for menadione. We noted the presence of two phenotypically (differences concerning antimicrobial susceptibility) and genetically distinct SCV strains in one patient, as well as the growth of two genetically related SCVs that differed in terms of their morphology and the type of auxotrophy in another. Seven out of eight SCVs were resistant to meticillin and gentamicin. In addition, antibiotic sensitivity testing revealed three multidrug-resistant SCV-normal-morphology isolate pairs. One S. epidermidis SCV harboured icaADBC genes and was found to be a proficient biofilm producer. This paper highlights the involvement of CNS SCVs in the aetiology of PJIs, including what is believed to be the first report of a S. warneri SCV. These subpopulations must be actively sought in the routine diagnosis of implant-associated infections. Moreover, in view of the phenotypic and genetic diversity of some SCV pairs, particular attention should be paid to the investigation of all types of observed colony morphologies, and isolates should be subjected to antimicrobial susceptibility testing.

Antimicrobial susceptibility and biofilm formation of Staphylococcus epidermidis small colony variants associated with prosthetic joint infection

We determined the frequency of isolation of non-aureus staphylococcal small colony variants (SCVs) from 31 patients with staphylococcal prosthetic joint infection (PJI) and described the antimicrobial susceptibility, auxotrophy, and biofilm-forming capacity of these SCVs. Eleven non-aureus SCVs were recovered, all of which were Staphylococcus epidermidis, and none of which was auxotrophic for hemin, menadione, or thymidine. Aminoglycoside resistance was detected in 5. Two were proficient, and 7 were poor, biofilm formers. With passage on antimicrobial free media, we observed a fluctuating phenotype in 3 isolates. We also noted a difference in antimicrobial susceptibility of different morphology isolates recovered from the same joints despite similar pulsed-field gel electrophoresis patterns. Our findings suggest S. epidermidis SCVs are common in PJI, and while they have a similar appearance to S. aureus SCVs, they do not necessarily share such characteristics as aminoglycoside resistance; auxotrophy for hemin, menadione, or thymidine; or enhanced biofilm formation. We also underscore the importance of antimicrobial susceptibility testing of all morphologies of isolates recovered from PJI.

Application of the antibiotic batumin for accurate and rapid identification of staphylococcal small colony variants

BackgroundStaphylococcus aureus is a major human pathogen causing significant morbidity and mortality. The S. aureus colonies in osteomyelitis, in patients with cystic fibrosis and patients with endoprosthesis rejection frequently have an atypical morphology, i.e. staphylococcal small-colony variants, which form a naturally occurring subpopulation of clinically important staphylococci. Identification of these small colony variants is difficult, because of the loss of typical phenotypic characteristics of these variants.We wanted to improve and simplify the diagnosis of staphylococcal infection using a diagnostic preparation, consisting of 5 μg batumin paper disks. Batumin possesses a unique selective activity against all studied Staphylococcus spp., whereas all other species tested thus far are batumin resistant. We assessed the efficacy of the batumin diagnostic preparation to identify staphylococcal small colony variants, isolated from osteomyelitis patients.FindingsWith the batumin diagnostic preparation, all 30 tested staphylococcal small-colony variants had a growth inhibition zone around the disk of minimum 25 mm, accordant with the inhibition zones of the parent strains, isolated from the same patients.ConclusionsThe batumin diagnostic preparation correctly identified the small-colony variants of S. aureus, S. haemolyticus and S. epidermidis as belonging to the genus Staphylococcus, which differ profoundly from parental strains and are difficult to identify with standard methods. Identification of staphylococcal small-colony variants with the batumin diagnostic preparation is technically simple and can facilitate practical laboratory work.

High-Dose Daptomycin for Cardiac Implantable Electronic Device-Related Infective Endocarditis Caused by Staphylococcal Small-Colony Variants

High-Dose Daptomycin for Cardiac Implantable Electronic Device-Related Infective Endocarditis Caused by Staphylococcal Small-Colony Variants

Comparative in vitro activity of finafloxacin against staphylococci displaying normal and small colony variant phenotypes

Staphylococcal small colony variants (SCVs) are associated with chronic and relapsing infections and their intracellular location may shield them from host defences and antibiotics. Finafloxacin is a novel fluoroquinolone that exhibits optimal activity at slightly acidic conditions where the activity of other marketed fluoroquinolones decreases. Here, the in vitro activity of finafloxacin against clinical strain pairs consisting of an SCV and its clonally identical parental strain displaying the normal phenotype (NP) was compared with those of other fluoroquinolones at standard and low pH. In vitro activities of finafloxacin, ciprofloxacin, levofloxacin and moxifloxacin were tested against 28 methicillin-susceptible Staphylococcus aureus (MSSA) and three methicillin-resistant S. aureus (MRSA) SCV-NP strain pairs. Additionally, two S. aureus mutants (ΔhemB and ΔthyA) displaying the SCV phenotype and their wild-type strains as well as four SCV-NP pairs of coagulase-negative staphylococcal (CoNS) strains were included. MIC(50,) MIC(90) and MIC ranges were calculated based on MIC determination by Etest(®) at pH 5.8 and pH 7.2. Under acidic conditions, finafloxacin demonstrated superior activity against MSSA, MRSA and CoNS regardless of the phenotype. At neutral conditions, the activity against MSSA was as follows: moxifloxacin > finafloxacin > levofloxacin > ciprofloxacin. In comparison with methicillin-susceptible NP isolates, ciprofloxacin was less active against their corresponding SCVs. For other fluoroquinolones, there was no marked difference in activity against SCVs compared with NPs. Particularly in acidic body compartments, finafloxacin appears to be a promising new antibiotic for the treatment of persistent staphylococcal infections, including those caused by SCVs.

Small-colony Variants (SCVs) of Staphylococci: A Role in Foreign Body-associated Infections

Staphylococci have various strategies for resisting therapy that extend beyond classic mechanisms. Clinical experience with device-associated infections as well as with infections due to small-colony variants (SCVs) clearly shows that both antibacterial chemotherapy and host defense mechanisms are often unable to eliminate the pathogens and cure these infections. Of particular interest is the fact that in the past few years an increasing number of various foreign body-related infections due to staphylococcal SCVs have been reported. In this overview, the characteristics of SCVs recovered from clinical specimens and of defined mutants displaying the SCV phenotype are described. Their slow growth and changing biochemical and physiological features represent a challenge to clinical laboratory personnel, because recovery, identification, as well as susceptibility testing of these variants need particular efforts. In addition, the reduced susceptibility to aminoglycosides and the ability of SCVs to persist intracellularly require specific attention for the treatment of these infections. Thus, special efforts to search for these variants formed by Staphylococcus aureus or by coagulase-negative staphylococci should be considered when an infection is particularly resistant to therapy, persists for a long period or fails to respond to apparently adequate therapy with antimicrobial compounds.

Augmented Expression of Polysaccharide Intercellular Adhesin in a DefinedStaphylococcus epidermidisMutant with the Small-Colony-Variant Phenotype

While coagulase-negative staphylococci (CoNS), with their ability to form a thick, multilayered biofilm on foreign bodies, have been identified as the major cause of implant-associated infections, no data are available about biofilm formation by staphylococcal small-colony variants (SCVs). In the past years, a number of device-associated infections due to staphylococcal SCVs were described, among them, several pacemaker infections due to SCVs of CoNS auxotrophic to hemin. To test the characteristics of SCVs of CoNS, in particular, to study the ability of SCVs to form a biofilm on foreign bodies, we generated a stable mutant in electron transport by interrupting one of the hemin biosynthetic genes, hemB, in Staphylococcus epidermidis. In fact, this mutant displayed a stable SCV phenotype with tiny colonies showing strong adhesion to the agar surface. When the incubation time was extended to 48 h or a higher inoculum concentration was used, the mutant produced biofilm amounts on polystyrene similar to those produced by the parent strain. When grown under planktonic conditions, the mutant formed markedly larger cell clusters than the parental strain which were completely disintegrated by the specific beta-1,6-hexosaminidase dispersin B but were resistant to trypsin treatment. In a dot blot assay, the mutant expressed larger amounts of polysaccharide intercellular adhesin (PIA) than the parent strain. In conclusion, interrupting a hemin biosynthetic gene in S. epidermidis resulted in an SCV phenotype. Markedly larger cell clusters and the ability of the hemB mutant to form a biofilm are related to the augmented expression of PIA.

Staphylococcal small colony variants have novel mechanisms for antibiotic resistance.

Over the past 4 years, a variant subpopulation of Staphylococcus aureus has been characterized that is defective in electron transport. These organisms grow slowly and are typical of the previously described small colony variants (SCVs). Indeed, many earlier papers included data that are consistent with defective respiratory activity in SCVs. We present a hypothesis that serves as biochemical basis for the development of SCVs. These variants are particularly interesting because they have been associated with very persistent infections, and they are more resistant to many antibiotics than normal S. aureus. Because of their slow growth, atypical colonial morphology, and unusual biochemical profile, they are easily missed or misidentified in the clinical laboratory. This is of some significance, as this subpopulation is more resistant to antibiotics than the parent population from which they arose. When an infection is particularly resistant to therapy, persists for a long period, or fails to respond to apparently adequate antimicrobial therapy, clinicians and clinical laboratory personnel should consider special efforts to search for SCVs.