scholarly journals In vitro activity of methylene blue and EMB agar on colistin-resistant A.baumannii: An experimental study

2019 ◽  
Author(s):  
Deniz Gazel ◽  
Müşerref Tatman Otkun ◽  
Alper Akçalı
Keyword(s):  
Methylene Blue ◽  
Clinical Isolates ◽  
Colistin Resistance ◽  
Carbapenem Resistant ◽  
Atcc Strain ◽  
Mueller Hinton ◽  
Mueller Hinton Agar ◽  
Resistant Strains ◽  
Moderately Resistant

Abstract Background Colistin is one of the last resort antibiotics used against carbapenem-resistant Acinetobacter baumannii (AB); however, colistin resistance has been reported recently. Methylene blue (MB) is used in microbiology for staining, and in medicine as an antidote drug. Here, we investigated antimicrobial effects of MB and Eosin Methylene blue (EMB) agar against colistin-resistant AB strains. Methods The AB ATCC 19606 strain and 31 AB clinical isolates were included in the study. In the first round, ATCC strain and a clinical isolate were transformed into colistin-resistant forms, using Li's method, with increasing colistin concentrations. At each step, new MICs were determined and subcultures were inoculated to EMB and sheep blood agar (SBA). The colistin MIC values of the subcultures were also determined using Mueller Hinton Agar (MHA) containing 14 µg/mL MB. In the second round, colistin resistant clones of all collected clinical isolates (n=31) were obtained and screened to investigate their susceptibility to EMB agar by inoculating on SBA and EMB agar. Results At the beginning, the MICs of two strains were 0.5 µg/mL. At the last stage, both MICs had risen to 64 µg/mL. Subpopulations with high colistin resistance (>=32 µg/mL) were inhibited by MB and EMB agar, but could grow well on SBA. In MHA plates containing MB, the MICs decreased to the 0.5 µg/mL level for colistin-susceptible or moderately resistant clones. Additionally, clones with high colistin resistance showed atypical colony morphology on SBA. In the second round, MICs of the colistin resistant clones of all clinical isolates rose to 8 µg/mL after colistin exposure and 35% of those clinical isolates were inhibited by EMB agar while they could grow on SBA. Conclusion Highly resistant strains were totally inhibited by the effect of MB and EMB agar, while the MICs of the susceptible and moderate resistant clones decreased. EMB agar and MB may have inhibitory effects against colistin-resistant AB strains and MB may have a potential to be used as an antimicrobial drug. Secondly, using only EMB agar for subculturing may cause missing of colistin-resistant strains and giving incorrect identification or antibiogram reports in clinical microbiology laboratories.

scholarly journals 1455. Potential Mechanisms of Cefiderocol MIC Increase in Enterobacterales in In Vitro Resistance Acquisition Studies

2020 ◽  
Vol 7 (Supplement_1) ◽  
pp. S730-S730
Author(s):  
Yoshinori Yamano ◽  
Rio Nakamura ◽  
Miki Takemura ◽  
Roger Echols
Keyword(s):  
Iron Transport ◽  
Resistance Mechanisms ◽  
Altered Expression ◽  
Carbapenem Resistant ◽  
Mueller Hinton ◽  
Two Component ◽  
Mueller Hinton Agar ◽  
Related Proteins

Abstract Background Cefiderocol (CFDC) is a novel siderophore, iron-chelating cephalosporin, which is transported into bacteria via iron transporters. CFDC has potent in vitro and in vivo activity against all aerobic Gram-negative bacteria, including carbapenem-resistant strains. To date, clinical isolates with cefiderocol MIC >4 µg/mL have been found infrequently, in which the presence of a few β-lactamases or altered iron transport was found. We investigated potential new mechanisms causing CFDC MIC increases in non-clinical studies. Methods The mutation positions were determined by whole genome sequencing using four K. pneumoniae mutants including two KPC producers and one NDM producer that had shown CFDC MIC increases in previous in vitro resistance-acquisition studies. The mutant strains were obtained at the frequency of 10-7 to < 10-8 by spreading bacteria on standard Mueller‒Hinton agar medium containing CFDC at concentrations of 10× MIC, with or without apo-transferrin (20 μg/mL). CFDC MIC was determined by broth microdilution using iron-depleted cation-adjusted Mueller-Hinton broth based on Clinical and Laboratory Standards Institute guidelines. The emergence of MIC increase mutants was also assessed by in vitro chemostat models under humanized plasma pharmacokinetic exposures of CFDC. Results The possible resistance mechanisms were investigated. Mutation of baeS or envZ, sensors of two-component regulation systems, were found in three or two mutants among the tested four isolates, respectively, and caused the MIC to increase by 4–32-fold. The altered expression level of specific genes by the baeS or envZ mutation could affect CFDC susceptibility, but the specific genes have not been identified. In addition, the mutation of exbD, an accessory protein related to iron transport, was identified in one case and caused the MIC to increase by >8-fold. In vitro chemostat studies using two isolates (one NDM producer and one KPC producer) showed no resistance acquisition during 24-hour exposure. Table. Overview of mutation emergence in five isolates of K. pneumoniae Conclusion The mutation of two-component regulation systems (BaeSR and OmpR/EnvZ) and iron transport-related proteins were shown to be possible mechanisms causing CFDC MIC increases, but these mutants did not appear under human exposures. Disclosures Yoshinori Yamano, PhD, Shionogi & Co., Ltd. (Employee) Rio Nakamura, BSc, Shionogi & Co., Ltd. (Employee) Miki Takemura, MSc, Shionogi & Co., Ltd. (Employee) Roger Echols, MD, Shionogi Inc. (Consultant)

Evaluation of the in vitro activity of new polymyxin B analogue SPR206 against clinical MDR, colistin-resistant and tigecycline-resistant Gram-negative bacilli

2020 ◽  
Vol 75 (9) ◽  
pp. 2609-2615 ◽  
Author(s):  
Yawei Zhang ◽  
Chunjiang Zhao ◽  
Qi Wang ◽  
Xiaojuan Wang ◽  
Hongbin Chen ◽  
...  
Keyword(s):  
Stenotrophomonas Maltophilia ◽  
Enterobacter Cloacae ◽  
Polymyxin B ◽  
Vitro Activity ◽  
Clinical Isolates ◽  
In Vitro Activity ◽  
Gram Negative ◽  
Carbapenem Resistant ◽  
Resistant Strains

Abstract Background SPR206 is a novel polymyxin analogue. Activity against clinical isolates is little documented. Methods A collection of 200 MDR, carbapenem-resistant, tigecycline-resistant, colistin-resistant and non-MDR clinical isolates of Acinetobacter baumannii, Pseudomonas aeruginosa, Klebsiella pneumoniae, Escherichia coli, Enterobacter cloacae and Stenotrophomonas maltophilia was obtained from 50 centres across China (2016–17). All isolates were derived from respiratory tract, urine and blood samples. Strains were purposely selected on the basis of phenotypes, genotypes and specimen origins. MICs of SPR206 and other antimicrobials were determined. Results SPR206 was active against all bacteria tested except colistin-resistant isolates. The MIC50/90 values of SPR206 for colistin-resistant strains were comparable to known polymyxins (16/128 versus 8/128 mg/L). SPR206 exhibited potent activity against colistin-susceptible OXA-producing A. baumannii (MIC50/90 = 0.064/0.125 mg/L), NDM-producing Enterobacteriaceae (MIC50/90 = 0.125/0.25 mg/L) and KPC-2-producing Enterobacteriaceae (MIC50/90 = 0.125/0.5 mg/L). In fact, SPR206 was the most potent agent tested, with 2- to 4-fold lower MICs than colistin and polymyxin B for A. baumannii, P. aeruginosa and Enterobacteriaceae. Additionally, MIC values of SPR206 (MIC50/90 = 0.064/0.125 mg/L) were 16- to 32-fold lower than those of tigecycline (MIC50/90 = 2/2 mg/L) for tigecycline-susceptible carbapenem-resistant A. baumannii. Conclusions SPR206 showed good in vitro activity against MDR, tigecycline-resistant and non-MDR clinical isolates of Gram-negative pathogens. SPR206 also exhibited superior potency to colistin and polymyxin B, with 2- to 4-fold lower MIC50/90 values.

scholarly journals In vitro susceptibilities of Bordetella pertussis and Bordetella parapertussis to four fluoroquinolones (levofloxacin, d-ofloxacin, ofloxacin, and ciprofloxacin), cefpirome, and meropenem.

1996 ◽  
Vol 40 (3) ◽  
pp. 807-808 ◽  
Author(s):  
J E Hoppe ◽  
E Rahimi-Galougahi ◽  
G Seibert
Keyword(s):  
Bordetella Pertussis ◽  
Clinical Isolates ◽  
Bordetella Parapertussis ◽  
Mueller Hinton ◽  
Mueller Hinton Agar ◽  
Agar Dilution ◽  
Horse Blood

The in vitro activities of levofloxacin, ofloxacin, d-ofloxacin, ciprofloxacin, cefpirome, and meropenem against 34 clinical isolates each of Bordetella pertussis and Bordetella parapertussis were determined by agar dilution on Mueller-Hinton agar supplemented with 5% horse blood. Levofloxacin was as active as ciprofloxacin against both species (MIC, 0.06 microgram/ml) and more active than ofloxacin and d-ofloxacin. Cefpirome was more active against B. pertussis (MIC, 1.0 microgram/ml) than against B. parapertussis (MIC, > 2 micrograms/ml), while the reverse was true for meropenem (MIC, 2.0 micrograms/ml against B. pertussis and 1.0 microgram/ml against B. parapertussis).

scholarly journals 696. Mechanism of Cefiderocol high MIC mutants obtained in non-clinical FoR studies

2018 ◽  
Vol 5 (suppl_1) ◽  
pp. S251-S251 ◽  
Author(s):  
Akinobu Ito ◽  
Toru Nishikawa ◽  
Ryuta Ishii ◽  
Miho Kuroiwa ◽  
Yoshino Ishioka ◽  
...  
Keyword(s):  
Resistance Mechanisms ◽  
Cross Resistance ◽  
Rt Pcr ◽  
Carbapenem Resistant ◽  
Iron Transporter ◽  
Mueller Hinton ◽  
Sds Page ◽  
Mueller Hinton Agar ◽  
Resistant Strains ◽  
Overnight Culture

Abstract Background Cefiderocol (S-649266, CFDC) is a novel siderophore cephalosporin with activity against a wide variety of Gram-negative bacteria including carbapenem-resistant strains. We previously reported that CFDC is efficiently transported into Pseudomonas aeruginosa via iron transporter PiuA. In this study, we examined frequency of resistance of P. aeruginosa to CFDC, and investigated the resistance mechanisms of appeared colonies. Methods Frequency of resistance (FoR) was determined by plating an overnight culture of P. aeruginosa PAO1 on Mueller–Hinton Agar containing 4× or 10×MIC of CFDC or ceftazidime (CAZ). Appeared colonies were analyzed by whole-genome sequencing (WGS) to identify genomic mutations. The mRNA expression was determined by real-time RT-PCR, and pyoverdine production was determined by MALDI-TOF/MS and expression of outer membrane protein was analyzed by SDS–PAGE and proteomic analysis. Results The FoR to CFDC was 2.9 × 10–8 and <7.1 × 10–8, which were lower than those to CAZ (3.1 × 10–7 and 3.4 × 10–8) in the conditions of 4× and 10×MIC, respectively. MIC of CFDC against CFDC-derived mutant increased from 0.5 μg/mL (MIC against PAO1) to 2 μg/mL, and MICs of CAZ did not increase. In the case of CAZ-derived mutant, MICs of CAZ increased from 1 μg/mL (MIC against PAO1) to 16 μg/mL or higher, though MIC of CFDC did not increase, suggesting no cross-resistance between CFDC and CAZ. WGS identified mutations in upstream regions of pvdS (pvdS mutant), which regulates pyoverdine synthesis, or fecI (fecI mutant), which regulates the synthesis of iron transporter FecA contributing to the transport of iron citrate. The pvdS expression and pyoverdine production in the pvdS mutant were more than 4- and 6-fold higher than those in PAO1, respectively. The expression of fecA in the fecI mutant was more than ninefold higher than that in PAO1. Conclusion The MIC increase of CFDC against P. aeruginosa occurred due to the mutation of iron transporter-related genes. The resistance acquisition risks should be low as the frequency of resistance to CFDC was lower and the MIC increase of CFDC against the mutants was smaller than that of CAZ. In addition, no cross-resistance between CFDC and CAZ was observed. Disclosures A. Ito, Shionogi & Co., Ltd.: Employee, Salary. T. Nishikawa, Shionogi & Co., Ltd.: Employee, Salary. R. Ishii, Shionogi & Co., Ltd.: Employee, Salary. M. Kuroiwa, Shionogi & Co., Ltd.: Employee, Salary. Y. Ishioka, Shionogi & Co., Ltd.: Employee, Salary. N. Kurihara, Shionogi & Co., Ltd.: Employee, Salary. I. Sakikawa, Shionogi & Co., Ltd.: Employee, Salary. T. Ota, Shionogi & Co., Ltd.: Employee, Salary. M. Rokushima, Shionogi & Co., Ltd.: Employee, Salary. M. Tsuji, SHIONOGI & CO., LTD.: Employee, Salary. T. Sato, SHIONOGI & CO., LTD.: Employee, Salary. Y. Yamano, SHIONOGI & CO., LTD.: Employee, Salary.

scholarly journals In vitro Activity of Cefiderocol Against Gram-Negative Clinical Isolates Collected from Urinary Track Source: SIDERO-WT-2014/SIDERO-WT-2015

2017 ◽  
Vol 4 (suppl_1) ◽  
pp. S375-S376 ◽  
Author(s):  
Masakatsu Tsuji ◽  
Meredith Hackel ◽  
Roger Echols ◽  
Yoshinori Yamano ◽  
Dan Sahm
Keyword(s):  
Vitro Activity ◽  
Clinical Isolates ◽  
Gram Negative Bacteria ◽  
In Vitro Activity ◽  
Gram Negative ◽  
Carbapenem Resistant ◽  
Mueller Hinton ◽  
The Usa ◽  
Fermenting Bacteria

Abstract Background The global rise of carbapenem resistant Gram-negative bacteria such as carbapenem-resistant Enterobacteriaceae (CRE) and carbapenem-resistant non-fermenting bacteria is alarming and become threats to patient as only a few drugs remain active (e.g. colistin). Cefiderocol (S-649266) is a novel parenteral siderophore cephalosporin with potent activity against a wide variety of Gram-negative pathogens including carbapenem-resistant strains. This study evaluated the in vitro activity of cefiderocol and comparator agents against clinical isolates collected from urinary track source from North America. Methods A total of 3,323 Enterobacteriaceae, 263 Acinetobacter spp, 509 Pseudomonas aeruginosa, and 38 Stenotrophomonas maltophilia collected from the USA and Canada in 2014–2016 were tested. MIC was determined for cefiderocol, cefepime (FEP), ceftazidime-avibactam (CZA), ceftolozane-tazobactam (C/T), ciprofloxacin (CIP), colistin (CST), and meropenem (MEM) by broth microdilution and interpreted according to CLSI 2016 guidelines. All testing was done at IHMA, Inc. As recommended by CLSI, cefiderocol was tested in iron-depleted cation-adjusted Mueller Hinton broth. Based upon CLSI breakpoints, carbapenem-non-susceptible (CarbNS) strains were defined as follows: MEM: MIC ≥2 µg/mL for Enterobacteriaceae, ≥4 µg/mL for non-fermenters. Quality control testing was performed on each day of testing by using E. coli ATCC25922 and P. aeruginosa ATCC27853. Results Cefiderocol exhibited in vitro activity against 4,133 strains of Gram-negative bacteria with an overall MIC90 of 0.5 µg/mL. At 4 µg/mL cefiderocol inhibited the growth of 99.9% of the all isolates. MIC90 of cefiderocol against CarbNS Enterobacteriaceae was 4 µg/mL although MIC90 of other comparators were >64 or >8 (CST) µg/mL. The cefiderocol MIC90value was 1 µg/mL for CarbNS non-fermeneters. Conclusion Cefiderocol demonstrated potent in vitro activity against Enterobacteriaceae, A. baumannii, P. aeruginosa, and S. maltophilia isolates collected from a UTI source, with greater than 99.9% of isolates having MIC values ≤4 µg/mL. These findings indicate that this agent has high potential for treating cUTI infections caused by these problematic organisms, including isolates resistant to colistin. Disclosures M. Tsuji, Shionogi & Co.: Employee, Salary; M. Hackel, IHMA: Employee, Salary; R. Echols, Shionogi & CO., LTD: Consultant, Consulting fee; Y. Yamano, Shionogi & Co.: Employee, Salary

scholarly journals Efficacy of Colistin versus β-Lactams, Aminoglycosides, and Rifampin as Monotherapy in a Mouse Model of Pneumonia Caused by Multiresistant Acinetobacter baumannii

2002 ◽  
Vol 46 (6) ◽  
pp. 1946-1952 ◽  
Author(s):  
A. Montero ◽  
J. Ariza ◽  
X. Corbella ◽  
A. Doménech ◽  
C. Cabellos ◽  
...  
Keyword(s):  
Acinetobacter Baumannii ◽  
In Vivo Model ◽  
Carbapenem Resistant ◽  
Bactericidal Efficacy ◽  
Life Threatening ◽  
Resistant Strains ◽  
Immunocompetent Mice ◽  
Moderately Resistant

ABSTRACT The treatment of life-threatening infections due to carbapenem-resistant Acinetobacter baumannii has become a serious challenge for physicians worldwide. Often, only colistin shows in general good in vitro activity against these carbapenem-resistant strains, but its antibacterial efficacy in comparison with the antibiotics most used in clinical practice is not well known. We studied the efficacy of colistin versus those of imipenem, sulbactam, tobramycin, and rifampin in an experimental pneumonia model with immunocompetent mice. We used three strains of A. baumannii corresponding to the main clones (A, D, and E) involved in the outbreaks of our hospital, with different grades of resistance to imipenem (imipenem MICs of 1, 8, and 512 μg/ml, respectively) and to the other antibiotics. The MIC of colistin was 0.5 μg/ml for the three strains. Reduction of log10 CFU/g in lung bacterial counts, clearance of bacteremia, and survival versus results with controls were used as parameters of efficacy. Imipenem and sulbactam (Δlung counts: −5.38 and −4.64 log10 CFU/ml) showed the highest level of bactericidal efficacy in infections by susceptible and even intermediate strains. Tobramycin and rifampin (−4.16 and −5.15 log10 CFU/ml) provided good results against intermediate or moderately resistant strains, in agreement with killing curves and pharmacodynamics. On the contrary, colistin showed the weakest antibacterial effect among the antibiotics tested, both in killing curves and in the in vivo model (−2.39 log10 CFU/ml; P < 0.05). We conclude that colistin did not appear as a good option for treatment of patients with pneumonia due to carbapenem-resistant A. baumannii strains. Other alternatives, including combinations with rifampin, may offer better therapeutic profiles and thus should be studied.

scholarly journals 1280. In-Vitro Activity of Cefiderocol, Imipenem/Relebactam, & Ceftazidime/Avibactam in Ceftolozane/Tazobactam Resistant Strains of Multidrug Resistant Pseudomonas aeruginosa

2020 ◽  
Vol 7 (Supplement_1) ◽  
pp. S655-S655
Author(s):  
Daniel Navas ◽  
Angela Charles ◽  
Amy Carr ◽  
Jose Alexander
Keyword(s):  
Multidrug Resistant ◽  
Resistance Mechanisms ◽  
Vitro Activity ◽  
Clinical Isolates ◽  
Resistance Testing ◽  
Clinical Samples ◽  
In Vitro Activity ◽  
Carbapenemase Gene ◽  
Resistant Strains

Abstract Background The activity of imipenem/relebactam (I/R), ceftazidime/avibactam (CZA) and cefiderocol (FDC) were evaluated against clinical isolates of multidrug resistant (MDR) strains of P. aeruginosa which was resistant to ceftolozane/tazobactam (C/T). The recent increase of MDR P. aeruginosa strains isolated from clinical samples has prompted research and development of new antimicrobials that can withstand its multiple resistance mechanisms. C/T is an effective option for treatment of MDR P. aeruginosa in our facility with only 10% of resistance in MDR strains, but the emergence of resistance may occur due to the presence of a carbapenemase gene or an ampC mutation. Methods Antimicrobial susceptibility testing for C/T Etest® (bioMérieux, Inc.) were performed on all MDR strains initially screened by the VITEK2® (bioMérieux, Inc.). 10% (n=20) of all MDR isolates were resistant to C/T by the CLSI 2019 breakpoints. These resistant isolates were tested for presence of a carbapenemase gene using the GeneXpert CARBA-R (Cepheid®) PCR and against CZA Etest® (bioMérieux, Inc.) I/R gradient strips (Liofilchem®) and FDC broth microdilution (Thermo Scientific™ Sensititre™). Results A total of 20 clinical isolates of MDR P. aeruginosa resistant to C/T were tested following standardized CLSI protocols and techniques. All 20 isolates were screened for the presence of a carbapenemase gene (blaVIM, blaNDM, blaKPC, blaOXA-48, blaIMP). A blaVIM gene was detected in 6 (30%) out of 20 isolates. FDC demonstrated the greatest activity with 85% (n=17) of susceptible isolates (CLSI MIC &lt;4µg/dL). CZA (CLSI MIC &lt;8µg/dL) and I/R (FDA MIC &lt;2µg/dL) showed 15% (n=3) and 10% (n=2) of susceptible isolates respectively. FDC was active against all 6 blaVIM isolates, where all 6 strains were resistant to CZA and I/R as expected. 3 isolates tested non-susceptible against FDC; additional characterization was not performed at this time. Conclusion Based on these results, FDC demonstrated the greatest in-vitro activity against C/T resistant strains of MDR P. aeruginosa. FDC also demonstrated activity against all 6 MDR P. aeruginosa carrying blaVIM gene. FDC is a strong option to consider on MDR P. aeruginosa strains based on a resistance testing algorithm and a cost/effective protocol. Disclosures All Authors: No reported disclosures

scholarly journals In Vitro Activities of Daptomycin against 2,789 Clinical Isolates from 11 North American Medical Centers

2001 ◽  
Vol 45 (6) ◽  
pp. 1919-1922 ◽  
Author(s):  
Arthur L. Barry ◽  
Peter C. Fuchs ◽  
Steven D. Brown
Keyword(s):  
Clinical Isolates ◽  
In Vitro Activity ◽  
Broth Microdilution ◽  
In Vitro Susceptibility ◽  
Medical Centers ◽  
Mueller Hinton ◽  
Calcium Cations ◽  
Important Exception ◽  
Mueller Hinton Broth

ABSTRACT The in vitro activity of daptomycin is affected by the concentration of calcium cations in the test medium. Mueller-Hinton broth is currently adjusted to contain 10 to 12.5 mg of magnesium per liter and 20 to 25 mg of calcium per liter, but for testing of daptomycin, greater concentrations of calcium (50 mg/liter) are recommended to better resemble the normal concentration of ionized calcium in human serum. Two levels of calcium were used for broth microdilution tests of 2,789 recent clinical isolates of gram-positive bacterial pathogens. MICs of daptomycin were two- to fourfold lower when the broth contained additional calcium. For most species, however, the percentages of strains that were inhibited by 2.0 μg of daptomycin per ml were essentially identical with the two broth media. Enterococci were the important exception; i.e., 92% were inhibited when tested in calcium-supplemented broth but only 35% were inhibited by 2.0 μg/ml without the additional calcium. This type of information should be considered when selecting criteria for defining in vitro susceptibility to daptomycin.

Sign in / Sign up

Export Citation Format

Share Document