scholarly journals Involvement of Small Colony Variant-Related Heme Biosynthesis Genes in Staphylococcus aureus Persister Formation in vitro

2021 ◽  
Vol 12 ◽  
Author(s):  
Xuyang Wang ◽  
Weizheng Li ◽  
Wenjie Wang ◽  
Shiyong Wang ◽  
Tao Xu ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Acid Stress ◽  
Heme Biosynthesis ◽  
Rna Seq ◽  
Small Colony Variant ◽  
Persistent Infections ◽  
Allelic Replacement ◽  
Small Colony ◽  
The Relationship

Background: Persisters are important reasons for persistent infections, and they can lead to antibiotic treatment failure in patients and consequently chronic infection. Staphylococcus aureus small colony variants (SCVs) have been shown to be related to persistent infection. Mutations in the genes of the heme biosynthesis pathway lead to the formation of SCVs. However, the relationship between heme production genes and persister has not been tested.Methods:HemA and hemB were knocked out by allelic replacement from S. aureus strain USA500 separately, and then, the heme deficiency was complemented by overexpression of related genes and the addition of hemin. The stress-related persister assay was conducted. RNA-sequencing was performed to find genes and pathways involved in heme-related persister formation, and relative genes and operons were further knocked out and overexpressed to confirm their role in each process.Results: We found that heme biosynthesis deficiency can lead to decreased persister. After complementing the corresponding genes or hemin, the persister levels could be restored. RNA-seq on knockout strains showed that various metabolic pathways were influenced, such as energy metabolism, amino acid metabolism, carbohydrate metabolism, and membrane transport. Overexpression of epiF and operon asp23 could restore USA500∆hemA persister formation under acid stress. Knocking out operon arc in USA500∆hemA could further reduce USA500∆hemA persister formation under acid and oxidative stress.Conclusion: Heme synthesis has a role in S. aureus persister formation.

Phage Activity Against Planktonic and Biofilm Staphylococcus aureus Periprosthetic Joint Infection Isolates

2021 ◽  
Author(s):  
Katherine M. Caflisch ◽  
Robin Patel
Keyword(s):  
Staphylococcus Aureus ◽  
Successful Treatment ◽  
Periprosthetic Joint Infection ◽  
Joint Infection ◽  
Bacterial Isolates ◽  
Small Colony Variant ◽  
Small Colony ◽  
Planktonic State ◽  
Phage Activity

We recently reported the successful treatment of a case of periprosthetic joint infection (PJI) with phage. Phage activity against bacteria causing PJI has not been systematically evaluated. Here we examined the in vitro activity of seven lytic phages against 122 clinical isolates of Staphylococcus aureus recovered between April 1999 and February 2018 from subjects with PJI. Phages were assessed against planktonic and biofilm phenotypes. Activity of individual phages was demonstrated against up to 73% of bacterial isolates in the planktonic state and up to 100% of biofilms formed by isolates that were planktonically phage-susceptible. Susceptibility to phage was not correlated with small colony variant status. These results demonstrate that phages can infect S. aureus causing PJI in both planktonic and biofilm phenotypes, and thus are worthy of investigation as an alternative or addition to antibiotics in this setting.

scholarly journals Attenuated Vancomycin Bactericidal Activity against Staphylococcus aureus hemB Mutants Expressing the Small-Colony-Variant Phenotype

2008 ◽  
Vol 52 (4) ◽  
pp. 1533-1537 ◽  
Author(s):  
Brian T. Tsuji ◽  
Christof von Eiff ◽  
Pamela A. Kelchlin ◽  
Alan Forrest ◽  
Patrick F. Smith
Keyword(s):  
Staphylococcus Aureus ◽  
Bactericidal Activity ◽  
Pharmacodynamic Model ◽  
Small Colony Variant ◽  
Variant Phenotype ◽  
Small Colony ◽  
Bactericidal Activities

ABSTRACT The in vitro bactericidal activities of vancomycin against Staphylococcus aureus hemB mutants displaying the small-colony-variant phenotype and their parental strains were evaluated. Vancomycin killing activities against hemB mutants were markedly attenuated, demonstrating approximately 50% less effect, a result which was well described by a Hill-type pharmacodynamic model.

Antibiofilm and intraosteoblastic activities of rifamycins against Staphylococcus aureus: promising in vitro profile of rifabutin

2020 ◽  
Vol 75 (6) ◽  
pp. 1466-1473 ◽  
Author(s):  
Lélia Abad ◽  
Jérôme Josse ◽  
Jason Tasse ◽  
Sébastien Lustig ◽  
Tristan Ferry ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Biofilm Formation ◽  
Therapeutic Potential ◽  
Joint Infection ◽  
Small Colony Variant ◽  
Resistance Selection ◽  
Low Concentrations ◽  
Infection Models ◽  
Small Colony

Abstract Background Targeting biofilm-embedded and intraosteoblastic Staphylococcus aureus, rifampicin gained a pivotal role in bone and joint infection (BJI) treatment. Two other rifamycins, rifabutin and rifapentine, may represent better-tolerated alternatives, but their activity against bacterial reservoirs associated with BJI chronicity has never been evaluated. Objectives To evaluate the activities of rifampicin, rifabutin and rifapentine in osteoblast infection models. Methods Using three S. aureus isolates, rifamycins were compared regarding: (i) their intracellular activity in ‘acute’ (24 h) and ‘chronic’ (7 days) osteoblast infection models at 0.1× MIC, 1× MIC, 10× MIC and 100× MIC, while impacting infection-induced cytotoxicity (MTT assay), intracellular phenol-soluble modulin (PSM) secretion (RT–PCR), resistance selection and small colony variant (SCV) emergence; and (ii) their minimal biofilm eradication concentration (MBEC) and their MIC to prevent biofilm formation (bMIC). Results At 0.1× MIC, only rifabutin significantly reduced intracellular inoculum and PSM secretion. All rifamycins allowed a 50% reduction of intraosteoblastic inoculum at higher concentrations, with no difference between acute and chronic infection models, while reducing infection-induced cytotoxicity and PSM secretion. Dose-dependent emergence of intracellular SCVs was observed for all molecules. No intracellular emergence of resistance was detected. bMICs were equivalent for all molecules, but MBEC90s of rifapentine and rifabutin were 10- to 100-fold lower than those of rifampicin, respectively. Conclusions All rifamycins are efficient in reducing the S. aureus intraosteoblastic reservoir while limiting infection-induced cytotoxicity, with a higher activity of rifabutin at low concentrations. All molecules prevent biofilm formation, but only rifapentine and rifabutin consistently reduce formed biofilm-embedded bacteria for all isolates. The activity of rifabutin at lower doses highlights its therapeutic potential.

scholarly journals Reporter Metabolite Analysis of Transcriptional Profiles of a Staphylococcus aureus Strain with Normal Phenotype and Its Isogenic hemB Mutant Displaying the Small-Colony-Variant Phenotype

2006 ◽  
Vol 188 (22) ◽  
pp. 7765-7777 ◽  
Author(s):  
Jochen Seggewiß ◽  
Karsten Becker ◽  
Oliver Kotte ◽  
Martin Eisenacher ◽  
Mohammad Reza Khoschkhoi Yazdi ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Dna Microarrays ◽  
Capsular Polysaccharide ◽  
Arginine Deiminase ◽  
Aureus Strain ◽  
Small Colony Variant ◽  
Normal Phenotype ◽  
Persistent Infections ◽  
Genome Wide ◽  
Small Colony

ABSTRACT In this study, full-genome DNA microarrays based on the sequence of Staphylococcus aureus N315 were used to compare the transcriptome of a clinical S. aureus strain with a normal phenotype to that of its isogenic mutant with a stable small-colony-variant (SCV) phenotype (hemB::ermB). In addition to standard statistical analyses, systems biology advances were applied to identify reporter metabolites and to achieve a more detailed survey of genome-wide expression differences between the hemB mutant and its parental strain. Genes of enzymes involved in glycolytic and fermentative pathways were found to be up-regulated in the hemB mutant. Furthermore, our analyses allowed identification of additional differences between the normal-phenotype S. aureus and the SCV, most of which were related to metabolism. Profound differences were identified especially in purine biosynthesis as well as in arginine and proline metabolism. Of particular interest, a hypothetical gene of the Crp/Fnr family (SA2424) that is part of the arginine-deiminase (AD) pathway, whose homologue in Streptococcus suis is assumed to be involved in intracellular persistence, showed significantly increased transcription in the hemB mutant. The hemB mutant potentially uses the up-regulated AD pathway to produce ATP or (through ammonia production) to counteract the acidic environment that prevails intracellularly. Moreover, genes involved in capsular polysaccharide and cell wall synthesis were found to be significantly up-regulated in the hemB mutant and therefore potentially responsible for the changed cell morphology of SCVs. In conclusion, the identified differences may be responsible for the SCV phenotype and its association with chronic and persistent infections.

scholarly journals In VitroSusceptibility of ClinicalStaphylococcus aureusSmall-Colony Variants to β-Lactam and Non-β-Lactam Antibiotics

2018 ◽  
Vol 62 (4) ◽  
Author(s):  
Evgeny A. Idelevich ◽  
André Kriegeskorte ◽  
Nina Schleimer ◽  
Georg Peters ◽  
Christof von Eiff ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Prolonged Incubation ◽  
Content Type ◽  
Small Colony Variant ◽  
Normal Phenotype ◽  
Gradient Diffusion ◽  
Small Colony ◽  
Inhibition Zones

ABSTRACTTheStaphylococcus aureussmall-colony variant (SCV) phenotype has been associated with relapsing and antibiotic-refractory infections. However, little is known about the activities of antibiotics on clinical SCVs. Here, we demonstrated that SCVs without detectable auxotrophies were at least as susceptible to most β-lactam and non-β-lactam antibioticsin vitroas their corresponding clonally identical strains with a normal phenotype. After prolonged incubation, a regrowth phenomenon has been observed in gradient diffusion inhibition zones irrespective of the strains' phenotype.

scholarly journals In Vitro Activity of the Bacteriophage Endolysin HY-133 against Staphylococcus aureus Small-Colony Variants and Their Corresponding Wild Types

2019 ◽  
Vol 20 (3) ◽  
pp. 716 ◽  
Author(s):  
Nina Schleimer ◽  
Ursula Kaspar ◽  
Dennis Knaack ◽  
Christof von Eiff ◽  
Sonja Molinaro ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Treatment Options ◽  
Nasal Carriage ◽  
Resistance Mechanisms ◽  
Resistance Pattern ◽  
Small Colony Variant ◽  
Small Colony ◽  
Time Kill ◽  
Bacteriophage Endolysin

Nasal carriage of methicillin-susceptible (MSSA) and methicillin-resistant Staphylococcus aureus (MRSA) represents both a source and a risk factor for subsequent infections. However, existing MRSA decolonization strategies and antibiotic treatment options are hampered by the duration of administration and particularly by the emergence of resistance. Moreover, beyond classical resistance mechanisms, functional resistance as the formation of the small-colony variant (SCV) phenotype may also impair the course and treatment of S. aureus infections. For the recombinant bacteriophage endolysin HY-133, rapid bactericidal and highly selective in vitro activities against MSSA and MRSA has been shown. In order to assess the in vitro efficacy of HY-133 against the SCV phenotype, minimal inhibitory (MIC) and minimal bactericidal concentrations (MBC) were evaluated on clinical SCVs, their isogenic wild types, as well as on genetically derived and gentamicin-selected SCVs. For all strains and growth phases, HY-133 MIC and MBC ranged between 0.12 and 1 mg/L. Time-kill studies revealed a fast-acting bactericidal activity of HY-133 resulting in a ≥3 − log10 decrease in CFU/mL within 1 h compared to oxacillin, which required 4–24 h. Since the mode of action of HY-133 was independent of growth phase, resistance pattern, and phenotype, it is a promising candidate for future S. aureus decolonization strategies comprising rapid activity against phenotypic variants exhibiting functional resistance.

scholarly journals Comparative In Vitro Activity of Ceftobiprole against Staphylococci Displaying Normal and Small-Colony Variant Phenotypes

2005 ◽  
Vol 49 (10) ◽  
pp. 4372-4374 ◽  
Author(s):  
Christof von Eiff ◽  
Alexander W. Friedrich ◽  
Karsten Becker ◽  
Georg Peters
Keyword(s):  
Staphylococcus Aureus ◽  
Vitro Activity ◽  
Dilution Method ◽  
Agar Dilution Method ◽  
In Vitro Activity ◽  
Small Colony Variant ◽  
Antistaphylococcal Activity ◽  
Agar Dilution ◽  
Small Colony

ABSTRACT The antistaphylococcal activity of ceftobiprole was compared with those of cefuroxime, linezolid, and moxifloxacin by using the agar dilution method. Apart from three strains with small-colony variant phenotypes, all Staphylococcus aureus isolates tested were inhibited by ≤2 μg/ml of ceftobiprole. This compound exhibited an excellent antistaphylococcal activity, comparable to that of linezolid.

scholarly journals Extracellular DNA released by glycine-auxotrophic Staphylococcus epidermidis small colony variant facilitates catheter-related infections

2021 ◽  
Vol 4 (1) ◽  
Author(s):  
Junlan Liu ◽  
Zhen Shen ◽  
Jin Tang ◽  
Qian Huang ◽  
Ying Jian ◽  
...  
Keyword(s):  
Staphylococcus Epidermidis ◽  
Biofilm Formation ◽  
Underlying Mechanism ◽  
Extracellular Dna ◽  
Small Colony Variant ◽  
Persistent Infections ◽  
Small Colony

AbstractThough a definitive link between small colony variants (SCVs) and implant-related staphylococcal infections has been well-established, the specific underlying mechanism remains an ill-explored field. The present study analyzes the role SCVs play in catheter infection by performing genomic and metabolic analyses, as well as analyzing biofilm formation and impacts of glycine on growth and peptidoglycan-linking rate, on a clinically typical Staphylococcus epidermidis case harboring stable SCV, normal counterpart (NC) and nonstable SCV. Our findings reveal that S. epidermidis stable SCV carries mutations involved in various metabolic processes. Metabolome analyses demonstrate that two biosynthetic pathways are apparently disturbed in SCV. One is glycine biosynthesis, which contributes to remarkable glycine shortage, and supplementation of glycine restores growth and peptidoglycan-linking rate of SCV. The other is overflow of pyruvic acid and acetyl-CoA, leading to excessive acetate. SCV demonstrates higher biofilm-forming ability due to rapid autolysis and subsequent eDNA release. Despite a remarkable decline in cell viability, SCV can facilitate in vitro biofilm formation and in vivo survival of NC when co-infected with its normal counterparts. This work illustrates an intriguing strategy utilized by a glycine-auxotrophic clinical S. epidermidis SCV isolate to facilitate biofilm-related infections, and casts a new light on the role of SCV in persistent infections.

scholarly journals Increased Expression of Clumping Factor and Fibronectin-Binding Proteins by hemB Mutants of Staphylococcus aureus Expressing Small Colony Variant Phenotypes

2002 ◽  
Vol 70 (10) ◽  
pp. 5428-5437 ◽  
Author(s):  
Pierre Vaudaux ◽  
Patrice Francois ◽  
Carmelo Bisognano ◽  
William L. Kelley ◽  
Daniel P. Lew ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Surface Display ◽  
Human Embryonic Kidney Cells ◽  
Small Colony Variant ◽  
Reverse Transcription Pcr ◽  
Bacterial Adhesins ◽  
Transport Chain ◽  
Complete Restoration ◽  
Small Colony ◽  
Slow Growing

ABSTRACT Small colony variants (SCVs) of Staphylococcus aureus are slow-growing subpopulations that cause persistent and relapsing infections. The altered phenotype of SCV can arise from defects in menadione or hemin biosynthesis, which disrupt the electron transport chain and decrease ATP concentrations. With SCVs, virulence is altered by a decrease in exotoxin production and susceptibility to various antibiotics, allowing their intracellular survival. The expression of bacterial adhesins by SCVs is poorly documented. We tested fibrinogen- and fibronectin-mediated adhesion of a hemB mutant of S. aureus 8325-4 that is defective for hemin biosynthesis and exhibits a complete SCV phenotype. In this strain, adhesion to fibrinogen and fibronectin was significantly higher than that of its isogenic, normally growing parent and correlated with the increased surface display of these adhesins as assessed by flow cytometry. Real-time quantitative reverse transcription-PCR demonstrated increased expression of clfA and fnb genes by the hemB mutant compared to its isogenic parent. The influence of the hemB mutation on altered adhesin expression was confirmed by showing complete restoration of the wild-type adhesive phenotype in the hemB mutant, either by complementing with intact hemB or by supplementing the growth medium with hemin. Increased surface display of fibrinogen and fibronectin adhesins by the hemB mutation occurred independently from agr, a major regulatory locus of virulence factors in S. aureus. Both agr-positive and agr-lacking hemB mutants were also more efficiently internalized by human embryonic kidney cells than were their isogenic controls, presumably because of increased surface display of their fibronectin adhesins.

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