Elevated MIC Values Research Articles

Evaluation of the Activity of Triazoles Against Non-fumigatus Aspergillus and Cryptic Aspergillus Species Causing Invasive Infections Tested in the SENTRY Program.

The activity of isavuconazole and other triazoles against non-fumigatus (non-AFM) Aspergillus causing invasive aspergillosis was evaluated. A total of 390 non-AFM isolates were collected (1/patient) in 2017-2021 from 41 hospitals. Isolates were identified by matrix-assisted laser desorption ionization-time of flight mass spectrometry and/or internal spacer region/β-tubulin sequencing and tested by Clinical and Laboratory Standards Institute (CLSI) broth microdilution. CLSI epidemiological cutoff values were applied, where available. Isavuconazole showed activity against Aspergillus sections Flavi (n = 122; minimum inhibitory concentration [MIC]50/90, 0.5/1 mg/L), Terrei (n = 57; MIC50/90, 0.5/0.5 mg/L), Nidulantes (n = 34; MIC50/90, 0.12/0.25 mg/L), Versicolores (n = 7; MIC50, 1 mg/L), and Circumdati (n = 2; MIC range, 0.12-2 mg/L). Similar activity was displayed by other triazoles against those Aspergillus sections. Most of the isolates from Aspergillus sections Fumigati (n = 9), Nigri (n = 146), and Usti (n = 12) exhibited elevated MIC values to isavuconazole (MIC50/90, 2/-, 2/4, and 2/8 mg/L), voriconazole (MIC50/90, 2/-, 1/2, and 4/8 mg/L), itraconazole (MIC50/90, 2/-, 2/4, and 8/>8 mg/L), and posaconazole (MIC50/90, 0.5/-, 0.5/1, and >8/>8 mg/L), respectively. Isavuconazole was active (MIC values, ≤1 mg/L) against Aspergillus parasiticus, Aspergillus tamarii, Aspergillus nomius, Aspergillus nidulans, Aspergillus unguis, Aspergillus terreus, Aspergillus alabamensis, and Aspergillus hortai, while isavuconazole MIC values between 2 and 8 mg/L were observed against cryptic isolates from Aspergillus section Fumigati. Isavuconazole inhibited 96.1% of Aspergillus niger and 80.0% of Aspergillus tubingensis at ≤4 mg/L, the CLSI wild-type cutoff value for A niger. Voriconazole, itraconazole, and posaconazole showed similar activity to isavuconazole against most cryptic species. Isavuconazole exhibited potent in vitro activity against non-AFM; however, the activity of triazoles varies among and within cryptic species.

Staphylococcus aureus nasal colonization and susceptibility profile to antimicrobials in hemodialysis patients using a protocol of seven collections

BackgroundPatients colonized with Staphylococcus aureus in their nasal passages have a higher risk of acquiring infection, especially if they are immunocompromised or have comorbidities such as chronic renal failure undergoing hemodialysis (HD). ObjectiveThis study aimed to report the prevalence of nasal carriage of S. aureus among HD patients utilizing a seven-week sampling protocol and to assess the susceptibility of these isolates to various antimicrobial agents. MethodsOver seven consecutive weeks, nasal swab samples were collected from 47 HD patients, resulting in a total of 329 samples. The microorganisms were identified using biochemical methods and subjected to antimicrobial susceptibility testing via disk diffusion and microdilution techniques. ResultsOut of all the patients analyzed, 25 individuals (53.19%) were found to be colonized by S. aureus, with 21 of them displaying intermittent colonization. Additionally, 38% showed positive results for S. aureus in only the 6th or 7th week of sampling. Within the 58 isolates, 17.2% (n=10) exhibited methicillin (oxacillin)-resistance and 25.86% (n=15) displayed elevated vancomycin MIC values (2 µg/ml). Based on the results, daptomycin and gentamicin were found to be effective treatment options. However, 31% of the isolates (n=18) exhibited a MIC of 1 µg/ml for daptomycin. ConclusionOver half of the patients were colonized by S. aureus, but mostly on an intermittent basis. The identification of oxacillin resistance and high vancomycin and daptomycin MICs serve as warnings for possible future complications in managing bacteremia caused by S. aureus in these patients.

Evaluation of a Method for Standardized Antimicrobial Susceptibility Testing with Mycoplasma hyorhinis Field Isolates.

Organizations like the Clinical and Laboratory Standards Institute (CLSI) or the European Committee of Antimicrobial Susceptibility Testing (EUCAST) provide standardized methodologies for antimicrobial susceptibility testing of a wide range of nonfastidious and fastidious bacteria, but so far not for Mycoplasma spp. of animal origin. Recently, a proposed method for the standardized broth microdilution testing of Mycoplasma hyorhinis using commercial Sensititre microtiter plates was presented. In this study, we evaluated this broth microdilution method with 37 field isolates and tested their susceptibility toward the following antimicrobial agents: doxycycline, enrofloxacin, erythromycin, florfenicol, gentamicin, marbofloxacin, tetracycline, tiamulin, tilmicosin, tulathromycin, and tylosin. The isolates originated from different countries, isolation sites, and years. The broth microdilution method was carried out using a modified Friis broth as the culture and test medium. For macrolides and lincosamides, a bimodal distribution with elevated MIC values could be observed for almost half of the tested field isolates, deducing reduced susceptibility toward these substances. With a recently published protocol, we were able to test a variety of field isolates, and consistent data could be obtained. Using this method, monitoring studies of Mycoplasma hyorhinis isolates can be carried out in a comparable manner, and the observed susceptibility profiles can be screened for possible changes in MIC values in the future.

In Vitro Activity of Cefiderocol against a Global Collection of Carbapenem-Resistant Acinetobacter baumannii Isolates.

Background: Cefiderocol is a novel siderophore cephalosporin with potent activity against multi-drug-resistant Gram-negative pathogens including carbapenem-resistant Acinetobacter baumannii (CRAB). Methods: The susceptibility of 313 non-duplicate CRAB isolates with defined carbapenem resistance mechanisms from a global collection to cefiderocol, ceftazidime, ceftazidime/avibactam, ceftolozane/tazobactam, ciprofloxacin, colistin, imipenem/relebactam, meropenem, meropenem/vaborbactam, minocycline, and piperacillin/tazobactam was determined using the broth microdilution method. Isolates were obtained from various body sites from patients in 47 countries in five world regions between 2012 and 2016. The identification of carbapenem resistance mechanisms and assignment to A. baumannii international clonal lineages were based on whole genome sequencing. Results: Cefiderocol showed greater activity than comparator antimicrobials of the β-lactam class, including novel β-lactams combined with β-lactamase inhibitors, ciprofloxacin, and minocycline. Cefiderocol MIC50 and MIC90 values were 0.5 mg/L and 4 mg/L, respectively, while colistin had comparable activity with a higher MIC50 at 1 mg/L and a lower MIC90 value of 2 mg/L. Many isolates with elevated cefiderocol MICs ≥ 4 mg/L represented A. baumannii international clone (IC) 1 and harbored a metallo-β-lactamase. Conclusions: While cefiderocol is a useful addition to the limited armamentarium of drugs targeting this problematic pathogen, a considerable part of CRAB isolates had elevated MIC values in a range of 4 -> 32 mg/L, including all isolates with a metallo-β-lactamase.

Susceptibility patterns of amphotericin B, itraconazole, posaconazole, voriconazole and caspofungin for isolates causing invasive mould infections from the SENTRY Antifungal Surveillance Program (2018-2021) and application of single-site epidemiological cutoff values to evaluate amphotericin B activity.

We evaluated the activity of amphotericin B, itraconazole, posaconazole, voriconazole and caspofungin against 1468 invasive moulds collected worldwide from 2018 to 2021. Most (>92%) of the Aspergillus spp. isolates were wildtype (WT) to amphotericin B, caspofungin and the azoles. Azole-non-wildtype A. fumigatus rates were higher in Europe (9.5%) and North America (9.1%) than Latin America (0.0%; only 12 isolates) and the Asia-Pacific region (5.3%). Amphotericin B and caspofungin were active against azole-non-wildtype A. fumigatus isolates. Posaconazole and amphotericin B were the most active agents against the Mucorales. Among the less common moulds, several expressed a pan-azole-resistant phenotype; many of these species also showed elevated MIC values (MIC, >2 mg/L) for amphotericin B and caspofungin. Although most isolates of Aspergillus spp. remain WT to the azoles, azole resistance is increasing in both North America and Europe. Amphotericin B and caspofungin exhibit potentially useful activity against azole-resistant A. fumigatus.

Novel Non-Hot Spot Modification in Fks1 of Candida auris Confers Echinocandin Resistance.

We determined the echinocandin susceptibility and FKS1 genotypes of 13 clinical isolates of Candida auris that were recovered from 4 patients at a tertiary care center in Salvador, Brazil. Three isolates were categorized as echinocandin-resistant, and they harbored a novel FKS1 mutation that led to an amino acid change W691L located downstream from hot spot 1. When introduced to echinocandin-susceptible C. auris strains by CRISPR/Cas9, Fks1 W691L induced elevated MIC values to all echinocandins (anidulafungin, 16 to 32×; caspofungin, >64×; micafungin, >64×).

Genetic Organization of Acquired Antimicrobial Resistance Genes and Detection of Resistance-Mediating Mutations in a Gallibacterium anatis Isolate from a Calf Suffering from a Respiratory Tract Infection

Gallibacterium (G.) anatis isolates associated with respiratory diseases in calves and harboring acquired antimicrobial resistance genes have been described in Belgium. The aim of this study was to analyze the genetic organization of acquired resistance genes in the G. anatis isolate IMT49310 from a German calf suffering from a respiratory tract infection. The isolate was submitted to antimicrobial susceptibility testing, and a closed genome was obtained by a hybrid assembly of Illumina MiSeq short-reads and MinION long-reads. Isolate IMT49310 showed elevated MIC values for macrolides, aminoglycosides, florfenicol, tetracyclines, and trimethoprim/sulfamethoxazole. The acquired resistance genes catA1, floR, aadA1, aadB, aphA1, strA, tet(M), tet(B), erm(B), and sul2 were identified within three resistance gene regions in the genome, some of which were associated with IS elements, such as ISVsa5-like or IS15DII. Furthermore, nucleotide exchanges within the QRDRs of gyrA and parC, resulting in amino acid exchanges S83F and D87A in GyrA and S80I in ParC, were identified. Even if the role in the pathogenesis of respiratory tract infections in cattle needs to be further investigated, the identification of a G. anatis isolate with reduced susceptibility to regularly used antimicrobial agents in cases of fatal bovine respiratory tract infections is worrisome, and such isolates might also act as a reservoir for antimicrobial resistance genes.

592. Impact of Elevated MIC Values on Echinocandin Pharmacokinetic-Pharmacodynamic (PK-PD) Candida glabrata Target Attainment (TA)

592. Impact of Elevated MIC Values on Echinocandin Pharmacokinetic-Pharmacodynamic (PK-PD) <i>Candida glabrata</i> Target Attainment (TA)

Suitability of Methods to Determine Resistance to Biocidal Active Substances and Disinfectants—A Systematic Review

Biocide resistance is an increasing concern. However, it is currently unknown if an elevated MIC value for an isolate correlates with a lower log10 reduction in suspension tests or carrier tests. The aim of this review was therefore to evaluate if isolates with a suspected tolerance to a biocidal active substance reveal an elevated MIC value and an impaired efficacy in suspension tests and carrier tests. A Medline search was done on 6 July 2022 using the following terms: “resistance biocidal MIC suspension” (16 hits), “resistance biocidal MIC carrier” (22 hits), “resistance biocidal suspension carrier” (41 hits), “tolerance biocidal MIC suspension” (1 hit), “tolerance biocidal MIC carrier” (4 hits) and “tolerance biocidal suspension carrier” (3 hits). Studies were included when a tolerance or resistance to the biocidal active substance or disinfectant was suspected and at least two of the three endpoints were evaluated in parallel in comparison to the control isolates. In three out of five studies, the elevated MIC values did not correlate with an impaired bactericidal efficacy against the biocide-tolerant isolates. In three out of five studies, an impaired activity in the suspension tests was described that correlated with an impaired efficacy in the carrier tests (peracetic acid-tolerant K. pneumoniae and glutaraldehyde-tolerant M. chelonae; the two other studies did not allow a comparison. Overall, the results from the suspension tests and tests under practical conditions allowed to determine a clinically relevant resistance.

Identification of a Novel Ceftazidime-Avibactam-Resistant KPC-2 Variant, KPC-123, in Citrobacter koseri Following Ceftazidime-Avibactam Treatment.

This study reported the identification of a novel ceftazidime-avibactam-resistant KPC-2 variant, KPC-123, in a Citrobacter koseri isolated from a patient in a Chinese hospital following ceftazidime-avibactam treatment of infection caused by OXA-232-producing Klebsiella pneumoniae. This novel KPC-123 consisting of 302 amino acids differs from KPC-2 by two insertions after positions 179 (ins179_TY) and 270 (ins270_DDKHSEA), respectively. Conjugation and cloning experiments confirmed that KPC-123 was able to confer high-level resistance to ceftazidime and ceftazidime/avibactam (MICs of 128 mg/L and 64/4 mg/L, respectively) and elevated MIC values of cefotaxime, cefepime, and aztreonam (4 mg/L, 2 mg/L, and 4 mg/L, respectively) but retained susceptibility to carbapenems. Whole-genome sequencing and genomic analysis revealed that blaKPC−123 within the “ISKpn27-blaKPC-ISKpn6” structure was located on a 93,814-bp conjugative plasmid that was almost identical to a blaKPC−2-carrying plasmid harbored in a K. pneumoniae isolate from the same sampling site of the patient, suggesting the transfer and in vivo evolution of this blaKPC-carrying plasmid. Hence, active surveillance of ceftazidime/avibactam resistance and the underlying mechanisms, which may facilitate the prevention and control of the dissemination of resistance, is needed.

Development of a harmonized method for antimicrobial susceptibility testing of Bordetella avium using broth microdilution and detection of resistance genes.

In response to a request from the Clinical and Laboratory Standards Institute (CLSI), the objective of this study was to develop a harmonized method for broth microdilution susceptibility testing of Bordetella (B.) avium, the major causative agent of infectious coryza in poultry. To find a suitable test medium, growth curves with four epidemiologically unrelated B. avium isolates were created in cation-adjusted Mueller-Hinton broth (CAMHB), CAMHB+2.5% lysed horse blood and veterinary fastidious medium. All isolates showed good growth in CAMHB, therefore MIC values were determined using this medium and the homogeneity of the values was determined. An essential MIC agreement of 99.7% was calculated. Testing of a larger strain collection (n=49) for their susceptibility to 24 antimicrobials confirmed the suitability of the tested method and revealed some isolates with elevated MICs of florfenicol (n=1), streptomycin (n=2), tetracyclines (n=5), and trimethoprim/sulfamethoxazole (n=6). PCR assays detected the resistance genes aadA1, dfrB1, floR, sul1, sul2 and tet(A). The method used enables easy reading and a good reproducibility of MIC values for B. avium. Application of the tested method allows harmonized resistance testing of B. avium and identification of isolates with elevated MIC values.

Differences in Fosfomycin Resistance Mechanisms between Pseudomonas aeruginosa and Enterobacterales.

Multidrug-resistant (MDR) Pseudomonas aeruginosa presents a serious threat to public health due to its widespread resistance to numerous antibiotics. P. aeruginosa commonly causes nosocomial infections including urinary tract infections (UTI) which have become increasingly difficult to treat. The lack of effective therapeutic agents has renewed interest in fosfomycin, an old drug discovered in the 1960s and approved prior to the rigorous standards now required for drug approval. Fosfomycin has a unique structure and mechanism of action, making it a favorable therapeutic alternative for MDR pathogens that are resistant to other classes of antibiotics. The absence of susceptibility breakpoints for fosfomycin against P. aeruginosa limits its clinical use and interpretation due to extrapolation of breakpoints established for Escherichia coli or Enterobacterales without supporting evidence. Furthermore, fosfomycin use and efficacy for treatment of P. aeruginosa are also limited by both inherent and acquired resistance mechanisms. This narrative review provides an update on currently identified mechanisms of resistance to fosfomycin, with a focus on those mediated by P. aeruginosa such as peptidoglycan recycling enzymes, chromosomal Fos enzymes, and transporter mutation. Additional fosfomycin resistance mechanisms exhibited by Enterobacterales, including mutations in transporters and associated regulators, plasmid-mediated Fos enzymes, kinases, and murA modification, are also summarized and contrasted. These data highlight that different fosfomycin resistance mechanisms may be associated with elevated MIC values in P. aeruginosa compared to Enterobacterales, emphasizing that extrapolation of E. coli breakpoints to P. aeruginosa should be avoided.

1454. Plazomicin Activity against Enterobacterales Isolates Producing Extended-Spectrum β-Lactamases (ESBLs), Carbapenemases, and Aminoglycoside-Modifying Enzymes (AMEs) from United States (US) Hospitals

BackgroundLimited therapeutic options are available for the treatment of multidrug resistant (MDR) organisms. Plazomicin (PLZ) is an aminoglycoside developed to overcome common aminoglycoside-resistance mechanisms. We evaluated the activity of PLZ and comparators against Enterobacterales isolates collected in 2018-2019 carrying genes encoding ESBLs, carbapenemases, and AMEs.MethodsAmong 3,899 Enterobacterales isolates from US hospitals susceptibility (S) tested using reference broth microdilution method, 619 isolates from selected species displaying elevated MIC values for cephalosporins, carbapenems and/or resistance (CLSI criteria) to amikacin (AMK), gentamicin (GEN) and tobramycin (TOB) were submitted to whole genome sequencing for detection of resistance genes.ResultsMost isolates producing ESBLs (n= 418) carried blaCTX-M (n= 386). The activity of PLZ (99.3% susceptible [S]) was comparable to that of colistin and higher than other comparators against ESBL isolates (Figure). AMK inhibited 96.4% of the isolates and GEN and TOB inhibited 57.9% and 43.5%, respectively. Only 34 isolates produced carbapenemases, including 19 KPC-2, 10 KPC-3, 1 each with VIM-1, OXA-181, NDM-5 and KPC-2-like plus 1 isolate carrying the genes encoding NDM-1 and OXA-232. These isolates displayed higher resistance rates to comparators and only PLZ, and tigecycline inhibited >90% of these isolates. AMK and GEN inhibited 67.6% and 55.9% of these isolates, respectively. PLZ was active against 97.7% of isolates carrying AME genes (n= 306) that carried aac(6’)-Ib-cr (n =177), aac(3)-IIa (n = 159) and aac(3)-IId (n =81), among others. Most of these isolates were resistant to GEN and TOB (only 10.8-14.1% S), but 92.8% were S to AMK. Three K. pneumoniae isolates carried 16S rRNA methyltransferases, 1 armA (also harboring NDM-1) and 2 rmtB1.ConclusionThe activity of PLZ against Enterobacteriaceae isolates carrying AMEs, ESBLs, and carbapenemases was greater than the activity of other aminoglycosides tested and comparable to those of tigecycline and colistin against carbapenemase-producing organisms. Isolates carrying genes encoding ESBLs, AMEs and carbapenemases are usually MDR and PLZ had activity against these organisms collected in the US.Figure 1 DisclosuresMariana Castanheira, PhD, 1928 Diagnostics (Research Grant or Support)A. Menarini Industrie Farmaceutiche Riunite S.R.L. (Research Grant or Support)Allergan (Research Grant or Support)Allergan (Research Grant or Support)Amplyx Pharmaceuticals (Research Grant or Support)Cidara Therapeutics (Research Grant or Support)Cidara Therapeutics (Research Grant or Support)Cipla Ltd. (Research Grant or Support)Cipla Ltd. (Research Grant or Support)Fox Chase Chemical Diversity Center (Research Grant or Support)GlaxoSmithKline (Research Grant or Support)Melinta Therapeutics, Inc. (Research Grant or Support)Melinta Therapeutics, Inc. (Research Grant or Support)Melinta Therapeutics, Inc. (Research Grant or Support)Merck (Research Grant or Support)Merck (Research Grant or Support)Merck & Co, Inc. (Research Grant or Support)Merck & Co, Inc. (Research Grant or Support)Paratek Pharma, LLC (Research Grant or Support)Pfizer (Research Grant or Support)Qpex Biopharma (Research Grant or Support) Rodrigo E. Mendes, PhD, A. Menarini Industrie Farmaceutiche Riunite S.R.L. (Research Grant or Support)Allergan (Research Grant or Support)Allergan (Research Grant or Support)Basilea Pharmaceutica International, Ltd (Research Grant or Support)Cipla Ltd. (Research Grant or Support)Department of Health and Human Services (Research Grant or Support)GlaxoSmithKline (Research Grant or Support)Melinta Therapeutics, Inc. (Research Grant or Support)Merck (Research Grant or Support)Merck (Research Grant or Support)Pfizer (Research Grant or Support) Timothy B. Doyle, Allergan (Research Grant or Support)Allergan (Research Grant or Support)Cipla Ltd. (Research Grant or Support)Melinta Therapeutics, Inc. (Research Grant or Support)Pfizer (Research Grant or Support)Qpex Biopharma (Research Grant or Support) Valerie Kantro, n/a, Cipla Ltd. (Research Grant or Support) Helio S. Sader, MD, PhD, A. Menarini Industrie Farmaceutiche Riunite S.R.L. (Research Grant or Support)Allergan (Research Grant or Support)Allergan (Research Grant or Support)Allergan (Research Grant or Support)Cipla Ltd. (Research Grant or Support)Cipla Ltd. (Research Grant or Support)Melinta (Research Grant or Support)Merck (Research Grant or Support)Merck (Research Grant or Support)Paratek Pharma, LLC (Research Grant or Support)Pfizer (Research Grant or Support) Jaideep Gogtay, n/a, Cipla Ltd. (Employee) Sandhya Das, n/a, Cipla Ltd. (Research Grant or Support)

733. Ibrexafungerp Demonstrates Potent and Consistent In Vitro Activity Against &amp;gt;400 Global Candida auris Isolates, Including isolates with Elevated MIC’s to Echinocandins

Background Candida auris is an urgent global threat; a pathogen associated with high mortality (up to 60%), multi-drug resistance, the ability to spread from person-to-person and surface-to-person, presenting high risk for outbreaks in healthcare facilities. Echinocandins are the first-line treatment for patients with Candida auris infections given the high degree of resistance to azoles and polyenes. Ibrexafungerp is a novel IV/oral glucan synthase inhibitor (triterpenoid) antifungal with activity against Candida, Aspergillus and Pneumocystis spp, in Phase 3 development. We will present a compilation of >400 Candida auris isolates from four studies, including 32 Candida auris isolates with elevated MIC’s to the echinocandins.Methods In vitro MIC data for ibrexafungerp against Candida auris isolates were compiled from across 4 independent studies with the majority of isolates originating in the US and India. In vitro susceptibility was determined by broth micro-dilution using CLSI (M27-S3) and/or EUCAST methods. Overall, 445 isolates were evaluated including 32 isolates with elevated MIC values to one or more echinocandins.ResultsThe ibrexafungerp MIC90 value against the 445 clinical isolates was 1 μg/mL; the modal and MIC50 values were 0.5 μg/mL each. These results were consistent across the four studies and no differences were observed between MIC results generated using CLSI or EUCAST methods. Similar results were obtained for the 32 isolates with elevated MIC values to one or more of the echinocandins. Among this population, the mode, MIC50 and MIC90 were 0.5, 0.5, and 1 μg/mL, respectively. Only 1/32 of the echinocandin-resistant isolates had reduced sensitivity to ibrexafungerp (defined as > 2-dilutions vs the mode). This isolate was pan-resistant with elevated MICs to all three echinocandins (MICs = 4 μg/mL) as well as fluconazole (MIC >256 μg/mL) and amphotericin B (MIC = 1 μg/mL).ConclusionThis data demonstrates that ibrexafungerp possesses potent and consistent in vitro activity against Candida auris and remains highly active against C. auris isolates with high MIC’s to the echinocandins.DisclosuresKatyna Borroto-Esoda, PhD, SCYNEXIS, Inc. (Employee, Shareholder) David A. Angulo, MD, SCYNEXIS, Inc. (Employee, Shareholder) Nkechi Azie, MD, SCYNEXIS, Inc. (Employee, Shareholder)

Ceftazidime-avibactam activity against a challenge set of carbapenem-resistant Enterobacterales: Ompk36 L3 alterations and β-lactamases with ceftazidime hydrolytic activity lead to elevated MIC values

IntroductionThis study examined ceftazidime-avibactam activity against carbapenem-resistant Enterobacterales (CRE) clinical isolates and resistance mechanisms among non-metallo β-lactamase (MBL) producers displaying ceftazidime-avibactam MIC values at 4 mg/L. MethodsCRE isolates (286 of 8161 Enterobacterales) collected in Asia-Pacific, Europe and Latin America during 2016 were screened for carbapenemase genes. Selected isolates were susceptibility tested for ceftazidime-avibactam in the presence or absence of phenylalanine-arginyl β-naphthylamide (PAβN) and polymyxin B nonapeptide (PMBN). Genome sequences were investigated for the integrity of outer membrane protein (OMP) genes and multilocus sequence typing. qRT-PCR assays were conducted to determine expression of acrA, ampC, and OMP genes. ResultsCeftazidime-avibactam inhibited 99.2% of the Enterobacterales, 22 (78.7%) of the 286 CRE and 226 (100%) non-MBL producers. Among carbapenemase producers (85.3%; 244 of 286), the most common gene was blaKPC (76 blaKPC-3 and 46 blaKPC-2), followed by blaOXA-48-like (60 isolates) and blaNDM (37). Ceftazidime-avibactam MIC values at 4 mg/L were noted among 14 Klebsiella pneumoniae (13 carrying blaKPC and 1 blaCTX-M-15) mostly from Italy and Brazil and 1 Klebsiella aerogenes overexpressing ampC. PAβN did not significantly decrease ceftazidime-avibactam results, but adding PMBN did significantly decrease the MIC results for the combination. All K. pneumoniae isolates had a premature stop codon at OmpK35 and most isolates had L3 alterations of OmpK36, low expression of this gene, or OmpC disruption (K. aerogenes). Nine K. pneumoniae isolates belonged to clonal complex 258 and displayed intrahospital clonality. ConclusionCeftazidime-avibactam is an important addition to the armamentarium against multidrug-resistant organisms, and elevated MIC results for this combination seem to be associated with L3 OmpK36 alterations and β-lactamases able to hydrolyze ceftazidime.

In vitro synergy of echinocandins with triazoles against fluconazole-resistant Candida parapsilosis complex isolates

IntroductionCandida parapsilosis (C. parapsilosis) is a common non-albicans Candida species ranked as the second common cause of bloodstream infections. Azole resistance and elevated echinocandin MICs have been reported for these fungi. This study was conducted to determine the interactions between azoles and echinocandins against C. parapsilosis species complex. Materials and methodsFifteen fluconazole-resistant clinical isolates of C. parapsilosis complex were included: C. parapsilosis sensu stricto (n = 7), C. orthopsilosis (n = 5) and C. metapsilosis (n = 3). The activity of azoles (fluconazole, itraconazole) and echinocandins (anidulafungin, micafungin) alone and in combination was determined using checkerboard broth microdilution. The results were determined based on the fractional inhibitory concentration index (FICI). ResultsIn vitro combination of fluconazole with anidulafungin was found to be synergistic (FICI 0.07–0.37) and decreased the MIC range from 4–64 μg/mL to 0.5–16 μg/mL for fluconazole and from 2–8 μg/mL to 0.125–1 μg/mL for anidulafungin. Similarly, interactions of fluconazole with micafungin (FICI 0.25–0.5), itraconazole with anidulafungin (FICI 0.15–0.37) and itraconazole with micafungin (FICI 0.09–0.37) were synergistic. ConclusionThe combination of fluconazole and itraconazole with either anidulafungin or micafungin demonstrated synergistic interactions against C. parapsilosis species complex, especially against isolates with elevated MIC values. However, the use of these combinations in clinical practice and the clinical relevance of in vitro combination results remain unclear.

Corrigendum: Comparison of 21-Plex PCR and API 20C AUX, MALDI-TOF MS, and rDNA Sequencing for a Wide Range of Clinically Isolated Yeast Species: Improved Identification by Combining 21-Plex PCR and API 20C AUX as an Alternative Strategy for Developing Countries

[This corrects the article DOI: 10.3389/fcimb.2019.00021.].

Postsurgical Cataract Prophylaxis With Intravitreal "Triamcinolone-Moxifloxacin" May Not Be Optimal For Preventing Endophthalmitis.

Intravitreal injections of antibiotics and anti-inflammatories are used by some cataract surgeons for surgical prophylaxis. To support this prophylaxis, intravitreal triamcinolone-moxifloxacin (TM) and triamcinolone-moxifloxacin-vancomycin (TMV) were tested for preventing Staphylococcus aureus (SA) endophthalmitis in rabbits. Trademark formulations of TM (15/1 mg/mL) and TMV (15/1/10 mg/mL) were intravitreally injected into seven groups of eight rabbits each (A-G). Before intravitreal injection, the vitreous was first challenged with clinical SA endophthalmitis isolates (5,000 colony-forming unit) with varying minimum inhibitory concentrations (MICs in μg/mL) to moxifloxacin (denoted by the MIC at the end of each group listed): A) TMV-10, B) TM-10, C) Saline-10, D) TM-2, E) Saline-2, F) TM-0.032, and G) Saline-0.032. After 24 hr, the rabbit eyes were graded for clinical endophthalmitis and cultured for viable SA. Rabbits treated with TMV and challenged by SA with a moxifloxacin MIC of 10 μg/mL did not present with endophthalmitis (0/8, no eyes with endophthalmitis). For SA with moxifloxacin MICs of 10.0 and 2.0 μg/mL, TM did not prevent endophthalmitis (16/16, 100% of eyes with endophthalmitis). For SA with a moxifloxacin MIC of 0.032 μg/mL, endophthalmitis was prevented with TM (0/8, no eyes with endophthalmitis). All saline-treated eyes developed endophthalmitis (23/23, 100% of eyes with endophthalmitis). Intravitreal monotherapy with TM did not provide consistent prevention of SA endophthalmitis, whereas intravitreal TMV successfully prevented endophthalmitis because of SA with elevated MIC values to moxifloxacin. Cataract surgeons need to be aware that vancomycin seems to be essential for intravitreal prophylaxis to cover moxifloxacin resistance.

ISEcp1-mediated transposition of chromosome-borne blaCMY-2 into an endogenous ColE1-like plasmid in Escherichia coli.

BackgroundCMY-2 is the most prevalent pAmpC β-lactamase, but the chromosomal blaCMY-2 gene transfer via horizontal transmission has been seldom reported. This study aimed to describe an ISEcp1-mediated transposition of a chromosomal blaCMY-2 gene from Escherichia coli into a small endogenous ColE1-like plasmid, resulting in elevated resistance to extended-spectrum cephalosporins.MethodsThree ESCs-resistant ST641 E. coli strains EC6413, EC4103 and EC5106 harbored the blaCMY-2 gene. S1-PFGE, I-ceu I-PFGE, Southern blotting and electroporation experiments were performed to investigate the location and transferability of blaCMY-2. The genetic context and gene expression of blaCMY-2 in the original isolates and the corresponding electroporants were explored by PCR mapping, primer walking strategy and RT-qPCR.ResultsThe blaCMY-2-containing region (ISEcp1-blaCMY-2-∆blc-∆yggR-∆tnp1-orf7-orf8-orf9-∆tnp2-∆hsdR) was transposed into endogenous ColE1-like plasmid pSC137 in the process of electroporation at very low frequencies (10–8–10–9). The transpositions resulted in novel larger blaCMY-2-harboring ColE1-like plasmids with size of 14,845 bp, enabling increase in MICs of 2 to 8-fold for cefotaxime, ceftiofur, and ceftazidime in recipient strains over their respective original counterparts. Transcriptional level analysis revealed that the increased blaCMY-2 expression was correlated with elevated MIC values of cephalosporins. The blaCMY-2 transposition unit was identical to that in a clinical isolate E. coli TN44889 from France isolated in 2004.ConclusionsOur results firstly demonstrated that ISEcp1 mediated a transposition of chromosome-borne blaCMY-2 into an endogenous ColE1-like plasmid by electroporation. Amplification of the blaCMY-2 gene facilitates the strain adaptation to a changed environment with an elevated antibiotic pressure.

Susceptibility of Methicillin-Resistant and -Susceptible Staphylococcus aureus Isolates of Various Clonal Lineages from Germany to Eight Biocides.

Few studies have been conducted on the susceptibility of bacteria to biocides. A total of 182 methicillin-resistant and -susceptible Staphylococcus aureus isolates collected from healthy or diseased humans and animals in Germany were included in the present study. Sixty-three isolates of animal origin and 119 human isolates were tested for their MICs to eight biocides or heavy metals by the broth microdilution method. The MIC50 and MIC90 values of human and animal isolates were equal or differed by not more than 1 dilution step, and statistical analysis revealed that differences between MICs of human and animal isolates were not significant. However, when taking into account the multilocus sequence type (MLST), a strong tendency (P = 0.054) to higher MICs of silver nitrate was detected for clonal complex 398 (CC398) isolates from humans compared to those from animals. Furthermore, a comparison of MIC values from isolates belonging to different clonal lineages revealed that important human lineages such as CC22 and CC5 exhibited significantly (P < 0.05) higher MICs for the biocides chlorhexidine, benzethonium chloride, and acriflavine than the main animal lineage sequence type 398 (ST398). Isolates with elevated MIC values were tested for the presence of biocide and heavy metal tolerance-mediating genes by PCR assays, and the following genes were detected: mepA (n [no. of isolates containing the gene] = 44), lmrS (n = 36), norA (n = 35), sepA (n = 22), mco (n = 5), czrC (n = 3), smr (n = 2), copA (n = 1), qacA and/or -B (n = 1), qacG (n = 2), and qacJ (n = 1). However, only for some compounds was a correlation between the presence of a biocide tolerance gene and the level of MIC values detected.IMPORTANCE Biocides play an essential role in controlling the growth of microorganisms and the dissemination of nosocomial pathogens. In this study, we determined the susceptibility of methicillin-resistant and -susceptible S. aureus isolates from humans and animals to various biocides and heavy metal ions and analyzed differences in susceptibilities between important clonal lineages. In addition, the presence of biocide or heavy metal tolerance-mediating genes was investigated. We demonstrated that important human lineages such as CC22 and CC5 had significantly higher MIC values for chlorhexidine, benzethonium chloride, and acriflavine than the main farm animal lineage, ST398. In addition, it was shown that for some combinations of biocides and tolerance genes, significantly higher MICs were detected for carriers. These findings provide new insights into S. aureus biocide and heavy metal tolerance.