Methicillin-susceptible Staphylococcus Aureus Strains Research Articles

Antibiotic Efficacy against Methicillin-Susceptible Staphylococcus aureus Biofilms on Synthetic and Biological Vascular Grafts

To compare the efficacy of five antibiotics against methicillin-susceptible Staphylococcus aureus (MSSA) biofilms on the surface of four vascular grafts. In vitro study of two clinical MSSA strains (MSSA2 and MSSA6) and four vascular grafts (Dacron, Dacron-silver-triclosan, Omniflow-II, and bovine pericardium). After a 24-hour incubation period, the graft samples were divided into six groups: growth control (no treatment), ciprofloxacin 4.5mg/L, cloxacillin 100 mg/L, dalbavancin 300 mg/L, daptomycin 140 mg/L, and linezolid 20 mg/L. Quantitative cultures were obtained and results expressed as log10 colony-forming units per milliliter (CFU/mL). Analysis of variance was performed to compare biofilm formation between the different groups. Mean ± SD MSSA2 count on the growth control Dacron graft was 10.05 ± 0.31 CFU/mL. Antibiotic treatment achieved a mean reduction of 45%; ciprofloxacin was the most effective antibiotic (64%). Baseline MSSA2 counts were very low on the Dacron-silver-triclosan (0.50 ± 1.03 CFU/mL) and Omniflow-II (0.33 ± 0.78 CFU/mL) grafts. On the bovine pericardium patch, the count was 9.87 ± 0.50 CFU/mL, but this was reduced by a mean of 45% after antibiotic treatment (61% for ciprofloxacin). Mean MSSA6 count on the growth control Dacron graft was 9.63 ± 0.53 CFU/mL. Antibiotics achieved a mean reduction of 48%, with ciprofloxacin performing best (67% reduction). Baseline MSSA6 count on the Dacron-silver-triclosan graft was 8.54 ± 0.73 CFU/mL. Antibiotics reduced biofilm formation by 72%; cloxacillin was the most effective treatment (86%). The MSSA6 count on the untreated Omniflow-II graft was 1.17 ± 1.52 CFU/mL. For the bovine pericardium patch, it was 8.98 ± 0.67 CFU/mL. Mean reduction after antibiotic treatment was 46%, with cloxacillin achieving the greatest reduction (68%). In this in vitro study, ciprofloxacin and cloxacillin performed best at reducing biofilms formed by clinical MSSA strains on the surface of biological and synthetic vascular grafts.

1813. A Population-Based Multi-Year Assessment of the Incidence of High Inoculum Cefazolin (CZ) Resistance Amongst Methicillin-Susceptible Staphylococcus aureus (MSSA) Causing Bloodstream Infections (BSIs) over time in Calgary, Canada

Abstract Background Cefazolin (CZ) is a 1st-line therapy for methicillin-susceptible Staphylococcus aureus (MSSA) infections due to its tolerability, lower toxicity, and convenient dosing. However, some strains exhibit reduced susceptibility to CZ when present in high inoculum. Treatment failure and worsened outcomes are documented in BSI cohorts treated with CZ when infections are caused by strains demonstrating this inoculum effect (IE) (Miller, 2018). The objective of our study was to investigate the incidence of MSSA strains demonstrating IE over time across a general population, free of referral bias. Methods All initial isolates of first episodes of MSSA BSI from persons in Calgary, Canada were analyzed during 2012-14 and 2019. CZ minimum inhibitory concentration (MIC) was determined at standard (SI: 5x105 CFU/mL) and high inocula (HI: 5x107 CFU/mL) using the broth microdilution method at 24 hours as per 2009 CLSI criteria. Isolates were defined as having the CZ IE if they had a ≥ 4-fold difference between MIC at SI and HI. Isolates were defined as having a pronounced IE (pIE), if they were susceptible at SI but resistant at HI with an MIC ≥ 32µg/mL. Population density of Calgary was obtained from census data. Annual incidence rates (IR) were calculated by dividing the number of IE and pIE by the mid-year population data, and IR ratios (IRR) were calculated for IE and pIE by comparing 2019 to 2012 rates. Results During the years of study, 1040 individuals experienced at least one episode of MSSA BSI. The mean incidence rates of IE and pIE were 6.50 and 0.98 per 100,000 person-years, respectively. Comparing 2019 to 2012, rates of IE decreased significantly with an IRR of 0.67 (95% CI 0.47 – 0.95) but rates of pIE were not significantly different (IRR 1.24; 95% CI 0.43 – 3.86). Incidence rates of IE and pIE in Calgary, Canada in 2012-14 and 2019. MSSA BSIs in Calgary, Canada over the years are shown alongside corresponding rates of IE and pIE. The population density was used to find rates for IE/pIE per 100,000 person-years. Conclusion Population-based studies on MSSA BSI with the IE and pIE phenotype are lacking. Herein, we observed that while the incidence of MSSA with pIE has not changed, those occurring with the IE have decreased. Further studies on the clinical implications of pIE are needed. Disclosures All Authors: No reported disclosures.

Prevalence of a cefazolin inoculum effect associated with blaZ gene types, and clinical outcomes among methicillin-susceptible Staphylococcus aureus blood isolates of patients with infective endocarditis

ObjectivesA proportion of blaZ gene-positive methicillin-susceptible Staphylococcus aureus (MSSA) strains exhibits the cefazolin inoculum effect (CInE). Its clinical impact remains uncertain but could compromise the use of cefazolin in high-burden infections. To date, no study has been conducted in France or in Europe. We aimed to assess the prevalence of CInE and its association with blaZ beta-lactamase and S. aureus protein A (spa) types, and to assess the clinical outcomes in cefazolin-treated patients for infective endocarditis whose strain exhibited a CInE. MethodsThis was a French single-center retrospective study of 51 MSSA strains from patients of the Nantes endocarditis prospective cohort, conducted between 2013 and 2018. ResultsCefazolin MIC50 at high inoculum was 2 mg/L (IQR 1–2). CInE was found in 17.6 % of tested strains. Among blaZ-positive strains (n = 44), type A beta-lactamase was predominant (n = 25, 57 %). Thirty-seven S. aureus protein A (spa) types were found. No statistical association was shown between blaZ or spa types and CInE. CInE was neither associated with a higher rate of persistent bacteremia (25 % vs 56.3 %, p = 0.58) nor with clinical failure in patients treated with cefazolin, in comparison to patients with no CInE strain (25 % vs 56.3 %, p = 0.58). ConclusionThe cefazolin inoculum effect was found in a substantial number of Staphylococcus aureus strains; however, minimum inhibitory concentrations remained globally low. CInE was not associated with a higher proportion of clinical failure on treatment.

CHARACTERIZATION OF STAPHYLOCOCCUS AUREUS ISOLATED FROM BURN WOUND; STRONG ANTIBACTERIAL ACTIVITY OF PHAGE COCKTAIL AGAINST VANCOMYCIN INTERMEDIATE-RESISTANT STAPHYLOCOCCUS AUREUS

A universal problem is about spread of Staphylococcus aureus infections in burn patients. The present study aimed to learn about the molecular characteristics and the resistance pattern of S. aureus strains isolated from burn patients. In this cross-sectional study, we investigated 100 unique S. aureus isolated from burn patients by antimicrobial activity and biofilm formation and evaluated the effect of Complex Pyobacteriophage, a commercial bacteriophage cocktail, against the isolates mentioned above. Methicillin-resistant Staphylococcus aureus (MRSA) comprised 76%, and methicillin-susceptible Staphylococcus aureus (MSSA) comprised 24% of 100 S. aureus strains. The resistance rate among MRSA isolates was higher than compared of MSSA. Mupirocin resistance was found in 30% of isolates, with 28 (93.3%) and 2 (6.7%) strains of MRSA and MSSA, respectively, found. Vancomycin intermediate resistance in S. aureus (VISA) was 13% of MRSA strains. Two isolates were confirmed as vancomycin-resistant S. aureus (VRSA) strains and carried vanA. 31 and 62% of the total isolates showed inducible and constitutive resistance phenotypes. The rate of inducible and constitutive clindamycin resistance among MRSA strains was higher than MSSA strains. Biofilm production was detected in 66% of isolates. Strong, moderate, and weak producers accounted for 25%, 17%, and 24% of isolates. Phage analysis showed that 81% were susceptible to the phage cocktail, and only 19% were resistant to the phage cocktail. Our data indicated that VISA strains prevalence in the burn unit was mainly from S. aureus infections. Present work recommended that vancomycin treatment be closely monitored to prevent the spread of VISA and VRSA strains. Observations also highlighted the role of bacteriophage cocktails in eradicating S. aureus-related infections.

Comparison of Molecular Characteristics Between Methicillin-Resistant and -Susceptible Staphylococcus aureus Clinical Isolates by Whole-Genome Sequencing.

IntroductionThe transmission of methicillin-resistant Staphylococcus aureus (MRSA) and methicillin-susceptible Staphylococcus aureus (MSSA) are great public health concern worldwide. To better understand S. aureus evolution and dissemination, we compared the molecular features of MSSA and MRSA isolates.MethodsIn this study, 74 MSSA and 102 MRSA non-duplicate isolates were recovered from clinical samples between 2016 and 2020. Molecular epidemiology, antimicrobial resistance determinants, and virulence gene profiles were carried out by whole-genome sequencing (WGS).ResultsTwenty distinct sequence types were identified in MRSA isolates, with the most common being ST59, ST630, and ST338. The major genotypes of MSSA were ST188 and ST7. The toxin genes clfA, sek, and seq were significantly associated with MRSA, while splA/B, clfB, map, sdrC/D, and sem-sen-seo-seu were detected more frequently in MSSA isolates than MRSA (P < 0.05). The tst positive isolates were more commonly identified in CC1 and CC72, whereas lukE/D was mainly found in the CC7, CC15, CC88, and completely absent in CC59 clones.ConclusionOur results compared the genetic diversity between MRSA and MSSA strains, suggesting efforts to fight infections caused by MSSA need to be intensified due to MSSA isolates carrying wide range of virulence factors. Comparative epidemiological studies of large populations of MSSA and MRSA will be necessary in the future to understand how MSSA and MRSA populations may co-evolve and interact in the future.

Epidemiology and antimicrobial susceptibility of Staphylococcus aureus in children in a tertiary care pediatric hospital in Milan, Italy, 2017—2021

BackgroundMethicillin-resistant Staphylococcus aureus (MRSA) is highly prevalent worldwide and can cause severe diseases. MRSA is associated with other antibiotic resistance. COVID-19 pandemic increased antimicrobial resistance in adult patients. Only a few data report the antimicrobial susceptibility of S. aureus in the Italian pediatric population, before and during the COVID-19 pandemic.MethodsWe included all the S. aureus positive samples with an available antibiogram isolated from pediatric patients (< 18 years old) in a tertiary care hospital in Milan, Italy, from January 2017 to December 2021. We collected data on demographics, antimicrobial susceptibility, and clinical history. We compared methicillin-susceptible Staphylococcus aureus (MSSA) and MRSA strains. We calculated the frequency of isolation by year. The incidence of isolates during 2020 was compared with the average year isolation frequency using the univariate Poisson test. We compared the proportion of MRSA isolates during 2020 to the average proportion of other years with the Chi-squared test.ResultsOur dataset included a total of 255 S. aureus isolated from 226 patients, 120 (53%) males, and 106 (47%) females, with a median age of 3.4 years (IQR 0.8 – 10.5). The mean isolation frequency per year was 51. We observed a significant decrease of isolations during 2020 (p = 0.02), but after adjusting for the total number of hospitalization per year there was no evidence that the incidence changed. Seventy-six (30%) S. aureus were MRSA. Twenty (26%) MRSA vs 23 (13%) MSSA (p = 0.02) were hospital-acquired. MRSA strains showed higher resistance to cotrimoxazole, clindamycin, macrolides, levofloxacin, gentamicin, and tetracyclin than MSSA strains. None of MRSA were resistant to linezolid and vancomycin, one was resistant to daptomycin. The proportion of MRSA did not change during the COVID-19 pandemic. The overall clindamycin resistance was high (17%). Recent antibiotic therapy was related to MRSA infection.ConclusionThe proportion of MRSA did not change during the COVID-19 pandemic and remained high. Clindamycin should not be used as an empirical MRSA treatment due to its high resistance.

Ceftriaxone and methicillin-susceptible staphylococcus aureus: a perspective from pharmacokinetics/pharmacodynamics studies

ABSTRACT Introduction: Usage of ceftriaxone-based therapy to treat Methicillin-Susceptible Staphylococcus aureus (MSSA) infections is a controversial issue, from in vitro to clinical studies. Area covered: We conducted a literature review using PubMed of articles with ceftriaxone pharmacokinetics parameters and built a probability of target attainment (PTA) based on PK values from stable conditions (non-critically-ill patients) with goals of fT>55%, fT>75%, and fT>100%. Ceftriaxone’s minimal inhibitory concentration from 31 MSSA strains (0.25–64 mg/L) was used to build the cumulative fraction response (CFR). The isolates were clinically relevant from blood, bronchoalveolar lavage, and soft tissue biopsy. Expert opinion: The results from controversies about using ceftriaxone for MSSA infections have been commonly addressed in the literature. However, variables such as (i) pharmacokinetic profile, (ii) pharmacodynamic target, (iii) site of infection, and (iv) MIC distributions may influence divergences. From this pharmacokinetics-pharmacodynamics perspective, ceftriaxone may be a reasonable option for MSSA infections when the MIC50 and MIC90 were 4 mg/L and 8 mg/L. CFR analysis demonstrated that ceftriaxone 1 g q24 h could be used if bacteriostasis is the aim (fT>55%), while 1 g q12h should be used for bactericidal effects (fT>75% or fT>100%). These dosing regimens should be considered in other clinical trials.

Difference in Host Immune response to Methicillin-Resistant and Methicillin Sensitive Staphylococcus aureus (MRSA and MSSA) Endophthalmitis

ABSTRACT Purpose The study aimed to understand the differential immune response of methicillin susceptible Staphylococcus aureus (MSSA) and methicillin-resistant S. aureus (MRSA) strains in in vitro models of endophthalmitis. Methods Retinal pigment epithelium (RPE) and microglia cells (CHME-3) were exposed to MRSA and MSSA strains and analyzed for expression of inflammatory mediators by real-time quantitative PCR and validated by ELISA or immunofluorescence assay. Heatmap and STRING analysis was used to assess the differential immune expression. Results Both microglia and RPE expressed TLR-2, TLR-1, TLR-6, and TLR −9 after challenge with MRSA and MSSA strains though the expression varied. MRSA-infected cells induced higher expression of IL-1β, IL-8, 1 L-10, IL-6, and GM-CSF, while TNF-α and IFN-ϒ were downregulated in comparison to MSSA-infected cells. We also demonstrate that MRSA infection leads to increased activation of MMP-9 and MMP-2 in RPE cells, while microglia expressed only MMP-9 in MRSA-infected cells. Conclusions MRSA strain can induce an exacerbated immune response in retinal cells. Giving clues for potential targets in immunomodulatory therapies.

255. Ticagrelor Aids Platelet-Mediated Clearance in a Refractory Staphylococcus aureus Endovascular Infection with Septic Emboli

BackgroundThe pore-forming alpha-toxin produced by Staphylococcus aureus (SA) decreases the viability and increases the clearance of platelets, a critical element of innate immune defense in endovascular infection. Our group recently identified that ticagrelor (TICA) blocks alpha-toxin-induced platelet clearance, protecting mice in a lethal systemic SA infection model. Here, we describe a case report in which TICA, added to antimicrobial therapy of a persistent methicillin-susceptible SA (MSSA) bacteremia associated with a septic aortic thrombus, resulted in immediate bacteremia clearance. We further explore TICA synergy with antibiotics and human platelets in vitro.MethodsAntibiotic susceptibility of an MSSA strain from a patient treated with TICA for refractory bacteremia was tested by MIC and checkerboard assays in MHB or RPMI at standard (105 CFU/mL) or high (107 CFU/mL) inocula using TICA, ertapenem (ETP), cefazolin (CZ), or nafcillin (NAF) alone vs. ETP+CZ ± TICA. Killing assays with human platelets ± TICA against SA were also performed.ResultsSA bacteremia secondary to a septic aortic thrombus (>4 mm) with multiple secondary pyogenic foci refractory to standard CZ and subsequent salvage CZ+ETP for 5 days rapidly cleared within 24 h after the addition of TICA. Thrombocytopenia resolved concurrently. Discontinuation of TICA on day 12 led to rebound thrombocytopenia, and TICA was restarted, once again resulting in resolution of thrombocytopenia. TICA alone lacked in vitro activity against SA, nor was TICA synergistic with ETP+CZ. In contrast, addition of a physiological achievable concentration of TICA dramatically sensitized SA to human platelet killing (p< 0.001) in vitro.Figure 1 Figure 2 Figure 3 ConclusionIn a complex case of aortic plaque rupture with septic thrombus, multiple septic emboli, and refractory MSSA bacteremia, addition of TICA to antimicrobial therapy yielded unanticipated immediate clinical and microbiological success. The profound therapeutic effect of TICA in vivo was corroborated by the enhanced staphylocidal activity of human platelets in vitro in the presence of physiological concentrations of the antiplatelet agent. TICA warrants further study as adjunctive treatment of refractory SA bacteremia due to a primary endovascular focus when thrombocytopenia is present.Disclosures Victor Nizet, MD, Centauri Therapeutics (Advisor or Review Panel member)Cidara Therapeutics (Advisor or Review Panel member)InhibRx (Advisor or Review Panel member)Roche Pharmaceutical (Advisor or Review Panel member)

Effect of methicillin-resistant Staphylococcus aureus (MRSA) infection in human microglial cells during endophthalmitis

Background: Infection caused by methicillin-resistant Staphylococcus aureus, or MRSA, is a growing concern in medicine, and ophthalmology has not been spared. MRSA is the predominant cause of postoperative endophthalmitis which occurs as a sight-threatening complication of cataract surgery. Toll-like receptors (TLRs) and immune mediators are important segments of the host response to infection and are expressed and secreted by a variety of cells including human microglial cells. To understand how MRSA modulates the innate immune response in comparison to methicillin-susceptible Staphylococcus aureus (MSSA), causing a dysregulated host-induced pathophysiology in retinal microglial cells during endophthalmitis. Methods and materials: Clinical isolates of MSSA and MRSA strains from patients with endophthalmitis were used for infecting human microglial cells (CHME-3) at an MOI 10:1, following which the mRNA expression analysis for various TLRs, pro-inflammatory cytokines and anti-inflammatory cytokines, along with MMPs were assessed at regular time intervals until 24 hrs post-infection (p.i). The expression of IL-10, IL-6, IL-8, M-CSFs was further confirmed by ELISA and of TLR-4, IFN-γ, and IL-1β by Immunofluorescence assay. Results: MRSA had a higher growth rate compared to the susceptible strain from every time point p.i (p< 0.05). mRNA analysis showed that the MRSA strain had increased expression of IL-6, IL-8, IL-10, IL-1α, IL-1β, TNF-α, IFN-γ, MMP-2, MMP-9, TLR-1, TLR-2, TLR-6, TLR-9 in a time-dependent manner compared to susceptible strain. Increased protein level expression of IL-10, IL-6, IL-8, and GM-CSF were further confirmed in the culture supernatant, by ELISA. The expression of immune mediators (TLR-4, IL-1β, and IFN-γ) was assessed by immunostaining, which showed differences in immunofluorescence intensity in resistant stimulated versus susceptible stimulated human microglia cells. Conclusion: Our work demonstrates that microglial cells incite differential innate responses when infected with an MRSA strain, compared to the susceptible strain. It is also associated with the potential for substantial immunopathology, which underscores the need for tight control of inflammatory responses. These immune system interactions may result in enhanced bacterial proliferation but also provoke systemic cytokine responses associated with endophthalmitis.

Genetic analysis of methicillin-susceptible Staphylococcus aureus clinical isolates: High prevalence of multidrug-resistant ST239 with strong biofilm-production ability.

BackgroundThe distributions of methicillin‐susceptible Staphylococcus aureus (MSSA) are divers geographically with different genetic backgrounds. Data related to molecular characteristics of MSSA compare to methicillin‐resistant Staphylococcus aureus (MRSA) is sparse.MethodsIn this cross‐sectional study, antimicrobial susceptibility testing, virulence genes analysis, biofilm formation, accessory gene regulator (agr) typing, and multilocus sequence typing (MLST) characterized on 75 MSSA isolates.ResultsMultidrug‐resistance MSSA was found to be 84%. Forty‐eight (64%) isolates were toxinogenic with 34 and 14 isolates carrying pvl and tst representing 45.3% and 18.7%. The most common SE genes were sed (20%), sec (16%), and sea (16%). Fifty‐five (73.3%) isolates were confirmed as biofilm producer with a markedly high prevalence of fnbA (93.3%), fnbB (86.7%), icaA (65.3%), icaD (53.3%), can (24%), ebp (10.7%), and bap (1.3%). A total of 3 agr types (I, 73.3%; III, 16%; II, 10.7%) and 4 clonal complexes (CCs) and sequence types (STs), namely CC8/ST293 (45.3%), CC/ST22 (28%), CC/ST30 (16%), and CC/ST5 (10.7%) were detected in this study. All the high and low‐level mupirocin resistance strains belonged to ST239 and ST22 strains, respectively. All the fusidic acid‐resistant isolates carried fusC and belonged to ST30.ConclusionsThese findings indicated that ST239 with strong biofilm production ability is the most common type in MSSA strains isolated from patients. It seems that the antimicrobial resistance profiles, toxin, and biofilm formation were closely associated with specific STs. Further studies are required to identify and control of these clonal lineages in our area.

Molecular surveillance of methicillin-susceptible Staphylococcus aureus (MSSA) isolated over a one-year period from a Malaysian Teaching Hospital.

We report the results of a molecular surveillance study carried out on methicillin-susceptible Staphylococcus aureus (MSSA) isolated in a one-year duration from Hospital Canselor Tuanku Muhriz (HCTM), a tertiary hospital located in Kuala Lumpur, Malaysia. The first strain isolated from each MSSA infection in HCTM during the year 2009 was included into the study. Antimicrobial susceptibility testing and agr group typing were carried out for all strains; virulence gene (cna, seh, TSST-1 and PVL) typing results of the strains were obtained from a previous study. Pulsed-field gel electrophoresis (PFGE) was done on selected strains from the orthopedic ward. Relationship(s) between different typing methods used in the study was investigated, where a p value of <0.05 indicated significant association between typing methods. A total of 880 MSSA strains were included into the study. The strains were generally susceptible to most antibiotics, with most of them carrying cna and agr-I (51.6%, n=454; 39.8%, n=350, respectively). A total of 17 PFGE pulsotypes were identified using an 80% similarity cut-off value, where the main pulsotype (pulsotype E) consisted of 24 isolates (23.5%). agr-III strains were found to be usually positive for both cna and seh (p<0.05). In addition, some PFGE pulsotypes were also characteristic of certain virulence genes or agr groups. We did not identify a dominant MSSA clone circulating in HCTM in 2009. Nevertheless, results from this molecular surveillance will provide good baseline data for the hospital's second S. aureus surveillance planned for the year 2020.

Occurrence and Characteristics of Methicillin-Resistant and -Susceptible Staphylococcus aureus Isolated from the Beef Production Chain in Korea.

The emergence and persistence of methicillin-susceptible Staphylococcus aureus (MSSA) and methicillin-resistant S. aureus (MRSA) in livestock animals have been reported as a potential risk factor for transmission to humans. In this study, we investigated the nationwide prevalence and characteristics of MRSA and MSSA in the Korean beef production system, including retail markets, slaughterhouses, and cattle farms. From a total of 1,285 samples, only 5 MRSA strains were isolated: from a farmer (1 ST72 MRSA), a carcass sample from a slaughterhouse (1 ST72 MRSA), and beef cattle (3 ST5 MRSA). In addition, 11 MSSA strains were isolated from beef cattle (n=3), humans (1 farmer, 1 slaughterhouse worker, and 4 retail market workers), and carcass samples (n=1) and slaughterhouse environment (n=1). Although the prevalence of MRSA and MSSA in beef cattle was much lower than that reported in pigs, 5/5 MRSA and 2/11 MSSA strains displayed multiple drug resistance (MDR) phenotypes. Unlike the swine-associated MRSA, no correlation was found between tetracycline/zinc resistance and MDR phenotype. However, MRSA strains had an identical set of staphylococcal enterotoxins and exhibited enhanced levels of resistance to antimicrobial peptides (PMAP-36 and LL-37) compared to the MSSA strains. In conclusion, continued and systemic surveillance of livestock, meat products, and humans in close contact with livestock/meat products is necessary to prevent the transmission of MRSA and MSSA to humans.

Levonadifloxacin (WCK 771) exerts potent intracellular activity against Staphylococcus aureus in THP-1 monocytes at clinically relevant concentrations.

Objective . Levonadifloxacin is a broad-spectrum anti-staphylococcal drug that is under development. We investigated the in vitro activity of levonadifloxacin against methicillin-susceptible Staphylococcus aureus (MSSA) and methicillin-resistant S. aureus (MRSA) strains phagocytized in THP-1 monocytes to evaluate its scope for treatment of intracellular staphylococcal infections.Methods. The microdilution minimum inhibitory concentrations (MICs) of levonadifloxacin, moxifloxacin, levofloxacin and ciprofloxacin against MSSA ATCC 25923 and MRSA ATCC 43300 strains at pH 7.4±0.1 (original medium pH) and 5.5±0.1 (phagosome pH environment) were determined by following Clinical and Laboratory Standards Institute (CLSI) guidelines. The activity of antibiotics was investigated by extracellular and intracellular time-kill studies at 1-16× MIC concentrations. A suspension of ~5× log10 c.f.u. ml-1 test organism in supplemented RPMI 1640 medium was employed to determine the extracellular activity, while test organism phagocytized at a 4 : 1 ratio of bacteria to THP-1 monocytes was employed to investigate the intracellular activity. At intervals of 0, 2, 6 and 24 h, colony-forming unit (c.f.u.) counts were performed in triplicate on inoculated brain heart infusion (BHI) agar plates for both methods.Results. At pH 7.4, the MIC of levonadifloxacin against both tested S. aureus strains was 2, 8 and 16 times lower than those of moxifloxacin, levofloxacin and ciprofloxacin, respectively. At pH 5.5, the MIC of levonadifloxacin was reduced by ≥8× against both tested S. aureus strains compared to its MIC at pH 7.4. In contrast, comparator quinolones showed a fourfold elevation in MIC at pH 5.5. In the study assessing the extracellular bactericidal effect, levonadifloxacin at 1× MIC manifested ≥4.5 log10 c.f.u. ml-1 killing for both S. aureus strains. Moxifloxacin and levofloxacin also showed bactericidal activity, while ciprofloxacin showed no killing. In intracellular conditions, levonadifloxacin manifested 1.0 log10 and 2.0 log10 killing for intracellular S. aureus ATCC 25923 and ATCC 43300, respectively. These killing effects were better overall than those of comparator quinolones.Conclusions. Within a clinically achievable concentration range, levonadifloxacin achieved a 90-99 % intracellular reduction of MSSA and MRSA strains phagocytized in THP-1 monocytes. Therefore, levonadifloxacin has the potential to be a therapeutic option for the management of intracellular methicillin- and quinolone-resistant staphylococcal infections.

Complete Genome Sequence of GD1108, a Moderate-Virulence Strain of Human-Associated ST398 Methicillin-Susceptible Staphylococcus aureus.

Staphylococcus aureus multilocus sequence type 398 (ST398) is responsible for an increasing number of severe infections in humans. There are no reports detailing if all ST398 strains are equally virulent. We present the genome sequence of the moderate-virulence ST398 methicillin-susceptible Staphylococcus aureus strain GD1108, determined in a Caenorhabditis elegans infection model, to reveal the ST398 sublineage virulence.

Complete Genome Sequence of GD487, a High-Virulence Strain of Human-Associated ST398 Methicillin-Susceptible Staphylococcus aureus.

Multilocus sequence type 398 (ST398) methicillin-susceptible Staphylococcus aureus (MSSA) has been shown to have augmented pathogenicity in humans. However, it has not been determined whether all ST398 strains are equally virulent. We present here the genome sequence of a high-virulence ST398 MSSA strain, GD487, to explore potential insights into ST398 virulence.

Antibacterial effects of extracts obtained from plants of Argentina: Bioguided isolation of compounds from the anti-infectious medicinal plant Lepechinia meyenii

Ethnopharmacological relevanceThe mostly native species from Argentina are used in traditional medicine generally for the treatment of pain and inflammation, respiratory, gastro-intestinal and urinary disorders and as antiseptics. Aim of the studySince these ailments may be associated with bacterial infections and that it is necessary to discover alternative compounds with antibacterial activity, 69 extracts from these plants were screened for their activity against pathogenic bacteria. The most effective extract was then submitted to bioguided isolation to obtain the compounds responsible for this activity. Materials and methodsExtracts and fractions were screened using agar dilution, and compounds using microbroth dilution methods. A large panel of pathogenic bacteria was used, especially methicillin-susceptible Staphylococcus aureus (MSSA) and methicillin-resistant S. aureus (MRSA). Bioguided fractionation was performed using successive chromatographic techniques, while the chemical structures of the isolated compounds were determined by nuclear magnetic resonance (NMR). Additionally, a series of derivatives of the most active compound were prepared in order to study the chemical features required for achieving the antibacterial effect. ResultsLepechinia meyenii (Walp.) Epling (Lamiaceae) extract showed itself the most effective, with minimum inhibitory concentration (MIC) against Gram positive and negative bacteria ranging from 62.5 to 500 μg/mL, and showing better activity on MRSA than on MSSA. Activity-guided fractionation yielded the abietanes carnosol (1), rosmanol (2) and carnosic acid (3) as active principles, with MICs ranging from 15.6-31.2, 15.6–62.5 and 7.8–15.6 μg/mL, respectively against 15 MRSA strains, and 15.6–31.2, 31.2–62.5 and 7.8–15.6 μg/mL, respectively against 11 MSSA strains, maintaining higher activity against the resistant bacteria, as does the extract. In addition, Enterococcus faecalis was sensitive to 1–3 with MICs of 15.6–62.5 μg/mL. The structure activity analysis showed that 12-OH is necessary for remarkable activity, but methylation in C-20 significantly increased this, as observed with 20-methyl carnosate (5) displaying the greatest effect, even more so than 3, with MICs of 3.9 μg/mL against all the tested MRSA and 3.9–7.8 μg/mL against the MSSA. ConclusionsThe results of this study contribute to validate the traditional antibacterial use of species native to Argentina, particularly of L. meyenii. The chemical structures of the compounds obtained may aid the design of antibacterial agents, especially those effective against MRSA.

Clinical characteristics and prognosis analysis of patient with Methicillin-susceptible Staphylococcus aureus, or Methicillin-resistant Staphylococcus aureus Bacteremia

Objective To investigate the clinical characteristics and prognosis analysis of patient with bacteremia induced by Methicillin-susceptible Staphylococcus aureus (MSSA) or Methicillin-resistant Staphylococcus aureus (MRSA). Methods This retrospective study included 95 Staphylococcus aureus Bacteremia (SAB) patients admitted to Nanjing Drum Tower hospital from January 2012 to December 2017. According to the drug sensitivity tests, patients were divided into MSSA group and MRSA group. Clinical characteristics, systematic score and prognosis were compared between these two group. APACHE Ⅱ score and SOFA score were analyzed by independent-samples t test. The distribution of primary disease and basic disease, bacterium source, and prognostic factors were analyzed by Mann—Whitney U test or χ2 test. The cumulative survival was calculated by the Kaplan-Meier method. Wilcoxon rank sum test was selected for survival rate comparison. Results There are 56 MSSA strain and 39 MRSA strain among the 95 SAB patients, the detection rate is 58.95% and 41.05%, respectively. The most common primary disease of SAB is catheter-related infections(23 cases, 24.21%). Compared with MSSA group, the catheter-related infections has higher detection rate of MRSA obviously (P 0.05). Conclusions The majority of strains in the SAB patients is MSSA. In clinical characteristics, compared with MSSA, MRSA is more common in nosocomial infection, catheter related infection and renal dysfunction patients. There is no significant differences in prognosis between MSSA group and MRSA group. Key words: Staphylococcus aureus; Methicillin-resistant Staphylococcus aureus; Methicillin-susceptible Staphylococcus aureus; Bloodstream infection; Bacteremia; Clinical characteristic; Prognosis; Retrospective analysis

Genotyping of Antimicrobial Resistance and Virulence in Staphylococcus Isolated from Food of Animal Origin in Mexico.

Ninety-six methicillin-susceptible Staphylococcus aureus (MSSA) and 11 methicillin-resistant coagulase-negative staphylococci (MRCNS) were recovered from food of animal origin. Multi-drug resistance was detected in 34.1% of isolates. Tetracycline-resistant staphylococci harbored tetK gene (68.8%). Erythromycin/clindamycin-resistant staphylococci carried lnuA/lnuB genes frequently alone or combined with msrA gene. The sec gene was detected in 15.6% of MSSA and two isolates harbored the immune evasion cluster. The spa t337 predominated among MSSA strains. Two ermC-positive MRCNS isolates were observed, five mecA-positive carried SCCmec IVa and 6 were non-typeable by the IWG-SCC classification. These results demonstrate that food of animal origin can be a potential source for spreading of multidrug-resistance gene.

A large retrospective cohort study of cefazolin compared with flucloxacillin for methicillin-susceptible Staphylococcus aureus bacteraemia.

Antistaphylococcal penicillins (ASPs) are recommended as first-line treatment for invasive infections caused by methicillin-susceptible Staphylococcus aureus (MSSA). Cefazolin is an alternative option, but there is theoretical concern about its use as some MSSA strains produce beta-lactamases active against cefazolin. The study compared the outcomes in patients with MSSA infections treated with flucloxacillin and cefazolin. We analysed data from The Australia and New Zealand Co-operative Outcomes of Staphylococcal Sepsis (ANZCOSS) observational study, which included all consecutive unique episodes of Staphylococcus aureus bacteraemia from 27 hospital-based or independent microbiology laboratories from January 2007 to September 2013. In this retrospective analysis of prospectively collected data, we compared 30-day all-cause mortality in patients with MSSA bacteraemia treated with flucloxacillin to that in patients treated with cefazolin. We included data from 7312 episodes of MSSA bacteremia and found no difference in 30-day mortality in those treated with flucloxacillin (731/6520 [11.2%, 95% CI 10.9-12.5%]) compared to cefazolin (83/792 [10.7%, 95% CI 8.4-12.8%]), OR 0.93 (95% CI 0.72-1.17). In a propensity-adjusted analysis, mortality remained non-significantly lower in the cefazolin group (aOR 0.86 [95% CI 0.65-1.14]). This study supports the results from previous observational studies from other regions, while extending them to Australasia and to a much larger number of patients. Although this observational study indicates cefazolin is likely to have equivalent or superior outcomes to ASPs for MSSA bacteraemia, this can only be convincingly proven by a properly designed randomised controlled trial.