Nonsteroidal antiinflammatory drugs alter antibiotic susceptibility and expression of virulence-related genes and protein A of Staphylococcus aureus

Background/aim: Nonsteroidal antiinflammatory drugs (NSAIDs) including diclofenac, naproxen, ibuprofen, acetylsalicylic acid, and acetaminophen have been shown to have antimicrobial effects on various microorganisms. The aim of this study was to investigate the antibacterial effects of NSAIDs on Staphylococcus aureus. Materials and methods: Susceptibilities of S. aureus strains to NSAIDs with or without antimicrobials (moxifloxacin, vancomycin, ciprofloxacin, clindamycin, and gentamicin) were determined using the microdilution method and disk diffusion test. Expression levels of genes in the presence of drugs were investigated by real-time quantitative RT-PCR (qRT-PCR), and immunoblotting analysis was performed for staphylococcal protein A (SpA). Results: Our results showed that all NSAIDs were active against S. aureus strains with MIC values ranging from 195 µg/mL to 6250 µg/ mL. NSAIDs increased the antibiotic susceptibility of the strains, and diclofenac was found to be more effective than the other drugs. Drugs showed different effects on expression levels of virulence factor and/or regulatory genes. Immunoblotting analysis of SpA protein was mostly in accordance with qRT-PCR results. Conclusion: The regulatory/virulence factor genes and proteins of S. aureus investigated in this study may be reasonable targets for these drugs, and we suggest that the data may contribute to the field of infection control and antimicrobial resistance.

In vitro effects of ciprofloxacin, levofloxacin and moxifloxacin on Mycobacterium tuberculosis isolates.

INTRODUCTION: Increased tuberculosis prevalence, and isolation of multidrug resistant (MDR) Mycobacterium tuberculosis strains frequently as causative organisms from tuberculosis infections are resulted in increasing need of new anti-tuberculosis drugs. Nowadays, fluoroquinolones known to have fewer side effects than the other drugs used in treatment of tuberculosis are sometimes assessed even as first-line anti-tuberculosis drugs due to their in vitro and in vivo strong activity. It was aimed in this study to investigate phenotypically the fluoroquinolone susceptibility in MDR and non-MDR M. tuberculosis isolates. MATERIALS AND METHODS: A total of 126 MDR and non-MDR M. tuberculosis isolates from mycobacteriology laboratory of two hospitals in the Aegean Region of Turkey were included in the study. Ciprofloxacin (CIP), levofloxacin (LEV) and moxifloxacin (MXF) susceptibilities were assessed by agar proportion method according to the Clinical and Laboratory Standards Institute (CLSI) recommendations. RESULT: Twelve (15.2%), 5 (6.3%) and 4 (5.1%) of the MDR M. tuberculosis strains were resistant to CIP, LEV, MXF, respectively [resistance breakpoints (µg/mL); CIP (> 2), LEV (> 1), MXF (> 0.5)] while non-MDR strains were susceptible to CIP, LEV, MXF. CONCLUSIONS: Consequently, although high fluoroquinolone susceptibilities were evaluated as a pleasing data in this study, to preserve their efficiency for many years steadily, quinolone usage and resistance increment in MDR M. tuberculosis isolates should be monitored elaborately.

In vitro activity of linezolid against Mycobacterium tuberculosis strains isoalted from Western Turkey.

We investigated the linezolid susceptibility of Mycobacterium tuberculosis strains isolated from a tertiary care hospital in Izmir. A total of 67 M. tuberculosis strains (33 multidrug-resistant [MDR] and 34 non-MDR) were isolated and identified by the Tuberculosis Laboratory, Department of Microbiology and Clinical Microbiology, Faculty of Medicine, Ege University. The activity of linezolid was studied by the standard agar proportion method. For all of the strains, the MIC range was 0.06-1 mg/L, and the MIC(50) and MIC(90) values were 0.5 mg/L. No differences were observed between the MDR and non-MDR isolates. In general, linezolid was found to be effective for both the non-MDR and MDR M. tuberculosis strains.

[In vitro activities of quinupristin/dalfopristin in combination with vancomycin and gatifloxacin against Staphylococcus aureus and Enterococcus faecium clinical isolates]. / Staphylococcus aureus ve Enterococcus faecium klinik izolatlari üzerine kinupristin/dalfopristin ile vankomisin ve gatifloksasin kombinasyonlarinin in vitro aktiviteleri.

In this study, it was aimed to investigate the effects of quinupristin/dalfopristin in combination with vancomycin and gatifloxacin against Staphylococcus aureus and Enterococcus faecium isolates. A total 17 gram-positive bacterial isolates, composed of 4 methicillin-susceptible S. aureus (MSSA), 5 methicillin-resistant S. aureus (MRSA), 3 vancomycin-susceptible E. faecium (VSEF) and 5 vancomycin-resistant E. faecium (VREF) isolates, recovered from several clinical specimens in Ege University Faculty of Medicine, Turkey, were enrolled in this study. Antibiotic susceptibilities and interactions between antibiotics were determined by E-test (AB Biodisk, Sweden) method and fractional inhibitory concentration (FIC) indices were calculated for each combination. Synergistic activity was detected in only one MSSA isolate with the combination of quinupristin/dalfopristin and vancomycin (sigma FIC= 0.5). While the combination of quinupristin/dalfopristin and gatifloxacin yielded synergistic interaction in two MRSA and one MSSA isolate (sigma FIC= 0.37, 0.36 and 0.28, respectively) and additive interaction in one MSSA isolate (sigma FIC= 0.75), synergic activity was detected in one of the VREF isolate (sigma FIC= 0.29) and additive activity in two isolates (sigma FIC= 0.75 and 0.91, respectively). In this study, it was observed that the combination of quinupristin/dalfopristin and gatifloxacin was superior to the combination of quinupristin/dalfopristin and vancomycin especially in MRSA and VREF isolates. These in vitro results should be supported by in vivo studies which will guide the use of antibiotic combinations especially in the treatment of multi-resistant gram-positive bacterial infections.

[In vitro activity of linezolid/ertapenem combination in resistant gram-positive bacteria]. / Dirençli gram-pozitif bakterilerde linezolid/ertapenem kombinasyonunun in vitro aktivitesi.

The increasing prevalence of vancomycin-resistant enterococcus and methicillin-resistant staphylococcus infections has become a major therapeutic challenge and alternative therapy options are under consideration. In the present study, we aimed to evaluate the in vitro antibacterial activity of linezolid combined with ertapenem against two vancomycin-resistant Enterococcus faecium (VREF), two methicillin-resistant Staphylococcus aureus (MRSA) and two methicillin-resistant Staphylococcus epidermidis (MRSE) strains isolated from clinical specimens. In vitro activity of linezolid/ertapenem combination at 1/2 x MIC (minimal Inhibitory Concentration), 1 x MIC and 4 x MIC concentrations for each of the isolates was determined by time-kill curve method. At 1 x MIC and 4 x MIC concentrations, additive effect was detected for MRSA (at 6 and 24 h) and VREF (at 6 h) strains. Synergism was observed between two antibiotics at 4 x MIC concentration against one of the MRSE strains at 6th hour. Additive effect was determined at 6th and 24th hours in this strain at 1 x MIC concentration. No synergism was present in the other MRSE strain but additive interaction was detected at 6 h (1/2 x MIC) and 24 h (1 x MIC). Although these results support the use of linezolid/ertapenem combination in infections caused by resistant gram-positive strains, further in vitro and in vivo studies are necessary.

Quercetin inhibits swarming motility and activates biofilm production of Proteus mirabilis possibly by interacting with central regulators, metabolic status or active pump proteins.

BACKGROUND: Via its virulence factors such as swarm differentiation, biofilm and hemolysin production, urease enzyme, Proteus mirabilis causes urinary tract infections (UTIs), especially in complicated cases. Anti-pathogenic compounds attenuate the virulence of bacteria without showing 'cidal' activity and carry the potential to be used in the prevention and treatment of infectious diseases. PURPOSE: Search for anti-pathogenic effects of quercetin, which is a widely known and biologically active phytochemical, on Proteus mirabilis was the purpose of this study. In this context, the potential inhibitory activity of quercetin on swarming motility and biofilm production of a wild-type strain, P. mirabilis HI4320, was investigated in both phenotypically and genotypically. METHODS: Quercetin's effect on swarming motility was examined on LB agar plates, containing quercetin at various concentrations, by measuring the swarming diameter. The effect on biofilm formation, on the other hand, was analyzed by staining the formed biofilm of the bacterium, exposed to quercetin at various concentrations, with crystal violet and reading spectrophotometrically. Differences in expression levels of selected genes involved in swarming regulation were determined by real-time reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) to evaluate the mechanism of inhibitory action on swarming. Further investigations were carried out repeating swarming assays with the clones that derived from the wild-type strain by a TA system kit for direct one-step cloning and overexpressing the relevant genes. RESULTS: Our study revealed that quercetin inhibited swarming motility while activating biofilm production of P. mirabilis in direct proportion to the dose. Although all selected genes are inhibited in the same manner in liquid medium, and no significant differences could be detected in solid medium as demonstrated by RT-qPCR, experiments repeated with the clones overexpressing flhC (a component of flagellar transcriptional activator), speB (an agmatinase enzyme) and ompF (an outer membrane porin) genes showed that the respective clones could restore swarming, compensating for the inhibitory effect of quercetin. CONCLUSION: Quercetin's inhibitory effect on P. mirabilis swarming was possibly due to interactions with components of swarming regulators, the genes expressing polyamine coding enzymes that trigger swarm differentiation, or active pump proteins.

[Extended spectrum beta-lactamase types in Klebsiella pneumoniae strains isolated from blood cultures]. / Kan kültürlerinden izole edilen Klebsiella pneumoniae suslarinda genislemis spektrumlu beta-laktamaz varliginin arastirilmasi ve tiplendirilmesi.

The aim of this study was to investigate and type the extended spectrum beta-lactamases (ESBL) in Klebsiella pneumoniae strains isolated from blood cultures. Following the detection of antibiotic susceptibilities in 32 K. pneumoniae isolates, ESBL were detected in 13 (41%) of them by using double disc synergy test. Minimum inhibition concentrations for ceftazidime, cefotaxime, and aztreonam of ESBL positive strains were determined by E-test. After the extraction of the enzymes, the types of ESBLs were investigated by isoelectric focusing method. It was seen that, of all ESBL positive strains, one strain had four bands, one had three bands, six strains had two bands, and each of the others had only one beta-lactamase band. The results of polymerase chain reaction (PCR) revealed bla(SHV) in ten samples, bla(TEM) in six samples, and bla(SHV) with bla(TEM) in four samples. Ten SHV enzymes were typed as ESBL by PCR-RFLP (restriction fragment lenght polymorphism) method. The results of isoelectric focusing, PCR and RFLP performed in these ESBL positive K. pneumoniae isolates showed that the ESBL types could be SHV-2, SHV-5 and SHV-12 in the tested strains. It should always be taken into consideration that K. pneumoniae isolates could produce ESBLs and antibiotic treatment protocols should be adjusted in accordance.

The post-antibiotic effects of linezolid against Gram-positive pathogens.

OBJECTIVE: To investigate the post-antibiotic effect (PAE) of linezolid against methicillin-resistant and -susceptible staphylococci, vancomycin-resistant and -susceptible enterococci. METHODS: Clinical strains of Staphylococcus aureus (S. aureus), Staphylococcus epidermidis (S. epidermidis) and Enterococcus faecalis (E. faecalis) were isolated from hospitalized patients at Ege University Medical Faculty Hospital between June and September 2005. This study was made between September and December 2005. The PAE of linezolid was determined at the minimum inhibitory concentration (MIC) and 4 times the MIC concentrations for 60 minutes in Mueller-Hinton Broth (MHB). The duration of the PAE was obtained by following the recovery of bacterial growth in antibiotic free MHB measured colony forming units on Mueller-Hinton agar. RESULTS: All the straines were susceptible to linezolid. The PAE was greater at 4 times the MIC (0.5 - 2.4 hours) that at the MIC (0 - 1.7 hours) for linezolid against all organisms tested. The PAE for linezolid was slightly higher against E. faecalis strains than other organisms. CONCLUSION: In this study, it was demonstrated that linezolid had a moderate in vitro PAE against S. aureus, S. epidermidis and E.faecalis strains.

Emergence of phenotypic resistance to ciprofloxacin and levofloxacin in methicillin-resistant and methicillin-sensitive Staphylococcus aureus strains.

The emergence of phenotypic resistance to ciprofloxacin and levofloxacin in methicillin-sensitive and methicillin-resistant Staphylococcus aureus (MRSA) strains was studied. Twenty MRSA and 77 methicillin-sensitive S.aureus (MSSA) strains susceptible to both quinolones were investigated for resistance after single step or serial passages. No growth of 20 MRSA strains was observed at 4xMIC of levofloxacin after 48 h incubation, but 4 of 77 (5%) MSSA strains grew at the same concentration. At 4xMIC concentration of ciprofloxacin, 10 MSSA (13%) and five MRSA (25%) strains were grown. In the serial passages of MRSA strains, resistance to ciprofloxacin was 75 and 5% for levofloxacin by the third passage. In the seventh passage this resistance was 100 and 15%, respectively. In MSSA strains, resistance to ciprofloxacin was 75 and 19% to levofloxacin at the third passage and at the seventh passage, 100 and 61%, respectively. Emergence of ciprofloxacin resistance was more common and developed more rapidly than resistance to levofloxacin in both MRSA and MSSA strains.

Rational antibiotic use and academic staff.

This study was devised to determine the knowledge, attitude and behaviour of an educated group of people towards antibiotic use and self-medication with antibiotics. Of 1380 members of academic staff (excluding those from the Faculty of Medicine) of Ege University, 602 were chosen by systematic sampling methods. Two groups were formed. Group A included academic staff from the Faculties of Dentistry and Pharmacy and Group B, members of all other faculties. The mean age was 37.4+/-11.0 and 47.0% were females. The mean antibiotic knowledge score was 7.16+/-3.32. Self-medication with antibiotics was admitted by 45.8% of the total samle and 15.6% of the respondents used antibiotics until their symptoms disappeared regardless of the period of prescription. In Group A 48.8% and in Group B 80.7% of the respondents believed that antibiotics could be used for common cold. It is concluded that priority should be given to knowledge-based behaviour education programmes for the more highly educated community; there must also be restriction on the sale of antibiotics without prescription.

Prevalence of blaPER-1 and integrons in ceftazidime-resistant Gram-negative bacteria at a university hospital in Turkey.

In this study, we examined the prevalence of the PER-1 enzyme and the presence of clinically important integron classes in ceftazidime-resistant Gram-negative bacteria isolated at a university hospital. The blaPER-1 gene was detected by PCR in 33 (19.5%) of the total 169 Gram-negative bacteria, including 17 (23.3%) of the 73 Pseudomonas aeruginosa isolates and 16 (25%) of the 64 Acinetobacter baumannii complex isolates. Molecular fingerprinting revealed that blaPER-1 prevalence was mostly due to the dissemination of clonally related P. aeruginosa and A. baumannii complex strains. Class 1 integron (intI1) was detected in 52.7% of the 169 bacteria examined in this study. Its detection rates were estimated at 49.3% and 57.8% of the P. aeruginosa and A. baumannii complex strains isolated, respectively. It was also detected in 11 of the 16 Escherichia coli isolates and 5 of the 10 Klebsiella pneumoniae isolates. A single E. coli isolate and another K. pneumoniae isolate contained both class 1 and class 2 integrase genes. The results of this study revealed that clonally related blaPER-1-positive P. aeruginosa and A. baumannii complex strains, mostly harboring intI1, had disseminated at our hospital.

Irrational use of antibiotics among university students.

STUDY OBJECTIVE: The aim of the presented study was to evaluate the knowledge, attitude and behaviour of antibiotic usage in the student groups of a University in a country where the antibiotics are taken without prescriptions. DESIGN: Out of 5345 students (excluding those from the Faculty of Medicine) of Ege University, 678 were chosen by systematic sampling methods. The sampling group was divided into two groups. Group A included students from the Faculties of Pharmacy and Dentistry and Group B was composed of students of all other faculties. A questionnaire was used. MAIN RESULTS: The mean age was 21.0+/-3.0 and 58% were females. The aim of antibiotic use for common cold was 83.1% (P>0.05) and, to decrease fever was 32.1% (P<0.05) in both Groups. In Group A 36.1% and in Group B 44.9% of the respondents started antibiotics by themselves when they were ill (P>0.05) although 89.1% of both groups agree that antibiotics should be started with doctors' prescription. During their last infection in Group A 11.7% and in Group B 27.3% of the respondents used the same antibiotic as previously prescribed by their doctors and the use of antibiotics as advised by the doctors during their last infection was 50.8% in Group A and 35.3% in Group B. CONCLUSIONS: The use of antibiotics are found to be irrational among university students. National education programmes about the dangers of irrational antibiotic use and restriction of antibiotics without prescriptions should be the priority. This study indicated that knowledge regarding antibiotics cannot be evaluated alone since it did not always correlate with behaviour.