Different effects of sub-minimum inhibitory concentrations of gentamicin on the expression of genes involved in alginate production and biofilm formation of Pseudomonas aeruginosa

Background and Objectives: Antibiotics at sub-minimum inhibitory concentrations (sub-MIC) may alter bacterial viru- lence factors. The objective of this study was to investigate the effect of gentamicin at sub-MIC concentrations on the expres- sion of genes involved in alginate production and biofilm formation of Pseudomonas aeruginosa. Materials and Methods: The broth microdilution method was used to determine the MIC of gentamicin for three P. aeru- ginosa clinical isolates (P1-P3) and standard strains (PAO1 and 8821M). Alginate production and biofilm formation of the bacteria in the presence and absence of sub-MIC concentrations of gentamicin were measured using microtiter plate and carbazole assay, respectively. The real-time PCR method was used to determine the effect of gentamicin at sub-MIC con- centrations on the expression level of genes involved in biofilm formation (pelA and pslA) and alginate production (algD and algU). Results: Gentamicin at sub-MIC concentrations significantly reduced alginate production, biofilm formation, and the expres- sion of alginate and biofilm-encoding genes in clinical isolate P1. This inhibitory effect was also observed on the alginate production of 8821M strain and biofilm formation of PAO1strain. In clinical isolates, P2 and P3, alginate production, biofilm formation, and the expression of alginate and biofilm-encoding genes were significantly increased in exposure to sub-MIC concentrations of gentamicin. Conclusion: This study showed that different phenotypic changes in clinical isolates and standard strains of P. aeruginosa in exposure to sub-MIC concentrations of gentamicin are associated with changes in the expression of virulence genes. Further researches are required to understand the mechanisms involved in regulating the expression of virulence genes after exposure to sub-MIC concentrations of antibiotics.

Minimum Inhibitory Concentrations before and after Antibacterial Treatment in Patients with Mycobacterium abscessus Pulmonary Disease

The MICs of isolates from 86 patients with Mycobacterium abscessus (MABS); 46 with Mycobacterium abscessus subsp. abscessus (Mab), and 40 with Mycobacterium abscessus subsp. massiliense (Mma) were retrospectively analyzed. The main findings are as follows: i) Mma were significantly more susceptible to clarithromycin and azithromycin than Mab, and both subspecies tended to be more susceptible to clarithromycin than azithromycin. ii) Most isolates were susceptible to amikacin (93.0%), and over half to linezolid (55.8%). iii) Fifty-one isolates (59.3%) had MIC values of less than 1 μg/mL for sitafloxacin, and 65 (75.6%) had less than 0.5 μg/mL for clofazimine, which seems worth clinical investigating. iv) Among nine cases analyzed chronological changes, only two patients showed obvious MIC result changes even after the long-term multidrug treatment.

Genotypic Diversity of Ciprofloxacin Nonsusceptibility and Its Relationship with Minimum Inhibitory Concentrations in Nontyphoidal Salmonella Clinical Isolates in Taiwan

This study analyzed the genetic diversity of ciprofloxacin (CIP) nonsusceptibility and the relationship between two major mechanisms and minimum inhibitory concentrations (MICs) of CIP in nontyphoidal Salmonella (NTS). Chromosomal mutations in quinolone resistance-determining regions (QRDRs) and plasmid-mediated quinolone resistance (PMQR) genes were searched from ResFinder, ARG-ANNOT, and PubMed for designing the sequencing regions in gyrA, gyrB, parC, and parE, and the 13 polymerase chain reactions for PMQR genes. We found that QRDR mutations were detected in gyrA (82.1%), parC (59.0%), and parE (20.5%) but not in gyrB among the 39 isolates. Five of the 13 PMQR genes were identified, including oqxA (28.2%), oqxB (28.2%), qnrS (18.0%), aac(6′)-Ib-cr (10.3%), and qnrB (5.1%), which correlated with the MICs of CIP within 0.25–2 μg/mL, and it was found that oxqAB contributed more than qnr genes to increase the MICs. All the isolates contained either QRDR mutations (53.8%), PMQR genes (15.4%), or both (30.8%). QRDR mutations (84.6%) were more commonly detected than PMQR genes (46.2%). QRDR mutation numbers were significantly associated with MICs (p < 0.001). Double mutations in gyrA and parC determined high CIP resistance (MICs ≥ 4 μg/mL). PMQR genes contributed to intermediate to low CIP resistance (MICs 0.25–2 μg/mL), thus providing insights into mechanisms underlying CIP resistance.

1256. Clinical Response by Minimum Inhibitory Concentrations in Carbapenem-Resistant Pseudomonas aeruginosa Infections under Cefiderocol Compassionate Use Program

Background Cefiderocol (CFDC) has been developed for the treatment of serious infections caused by drug-resistant aerobic Gram-negative pathogens, including carbapenem-resistant (CR) Pseudomonas aeruginosa (CRPA). The current CFDC susceptibility breakpoints for P. aeruginosa differ between US Food and Drug Administration (FDA) and Clinical and Laboratory Standards Institute (CLSI) (Table). Data characterizing the impact of CFDC minimum inhibitory concentrations (MICs) on the clinical responses of patients treated with CFDC for CRPA are sparse. Methods We reviewed patients treated with compassionate-use CFDC (2 g, q8h or renally adjusted dosages) for infections caused by CRPA with no alternative treatment options. CFDC minimum inhibitory concentrations (MICs) were evaluated according to CLSI guidelines in iron-depleted cation-adjusted Müller–Hinton broth for available CRPA isolates. We then assessed physician-reported clinical responses to CFDC therapy and stratified results by CFDC MIC. Results There were 71 patients overall with CRPA treated with CFDC. Treatment duration ranged from 1 to 132 days. For the subset of 33 patients for whom CFDC MIC values were available, the most common infection sites were the respiratory tract (n=15), blood (n=12), and urinary tract (n=4). Patients could have had an infection at ≥1 sites and in other locations. CFDC MIC range was ≤0.03– &gt;64 µg/mL. The modal MIC value was 2 µg/mL (n=13; Table). CRPA isolates were susceptible to CFDC in 13/33 patients (39.4%) based on the FDA breakpoint (MIC ≤1 µg/mL) and in 31/33 patients (93.9%) based on the CLSI breakpoint (MIC ≤4 µg/mL). Clinical response was reported for 15/18 patients (83.3%) who had infections with CFDC MICs of 2–4 µg/mL, organisms that are considered susceptible by CLSI but not by FDA breakpoints (Table). Clinical response was reported in 6/13 patients (46.1%) with infections with CFDC MIC ≤1 µg/mL and in 1 of 2 patients (50.0%) with CFDC MIC ≥8 µg/mL (Table). 21 (63.6%) patients survived to Day 28 and there were no trends in mortality by CFDC MIC. Conclusion Clinical response rate was high for CRPA infections with CFDC MICs of 2–4 µg/mL, supporting the higher CLSI susceptibility breakpoint. Disclosures Michael J. Satlin, MD, MS, Achaogen (Consultant)Allergan (Research Grant or Support)BioFire Diagnostics (Research Grant or Support)Merck (Research Grant or Support)Shionogi (Consultant) David Fam, PharmD, Shionogi (Employee) Roger Echols, MD, Shionogi (Consultant) Christopher Longshaw, PhD, Shionogi (Employee) Miki Takemura, MS, SHIONOGI & CO., LTD. (Employee) Yoshinori Yamano, PhD, Shionogi (Employee)

1289. The Challenge of Treating Community-Associated Enterobacterales Infections in a Middle-Income Country: Data from SMART 2018-2019

Background Antimicrobial stewardship programs have been used widely in hospital settings due to the rise of resistant bacteria, antibiotic toxicities, and costs. Nevertheless, few efforts are done to prevent the rising antimicrobial resistance in community settings. The aim of our study was to evaluate the antimicrobial resistance from Enterobacterales community- and hospital-acquired infections in Southern Brazil. Methods A total of 272 Enterobacterales isolates (i.e., Escherichia coli, Klebsiella spp., Citrobacter spp., Enterobacter spp., Serratia spp., Proteus spp., and Providencia) were collected from 2018 and 2019. Broth microdilution method was used to determine minimum inhibitory concentrations for ceftriaxone, cefepime, levofloxacin, amikacin and ertapenem. Molecular evaluation of beta-lactamases (ESBLs, AmpC, and KPC) was also performed. Results Ninety-three, and a hundred and seventy-nine isolates were from community- and hospital-acquired infections, respectively. Similar MIC distribution was found between community and hospital settings (Table 1). Levofloxacin MIC of 8mg/L occurred in 38.7% (n=36) and 30.7% (n=55) of isolates from community- and hospital-acquired infections, respectively (Figure 1). Ceftriaxone MIC of 16mg/L occurred in 39.7%(n=37) and 39.1% (n=70) of isolates from community- and hospital-acquired infections, respectively (Figure 1). At last, cefepime MIC of 32mg/L occurred in 22% (n=21) and 25% (n=46) of isolates from community- and hospital-acquired infections, respectively. The following beta-lactamases were found in isolates from community-acquired group, ACT-MIR, CTX-M, SHV and TEM; while beta-lactamases from the hospital-acquired group were ACT-MIR, CMY II, KPC-2, CTX-M, SHV and TEM. Table 1. Enterobacterales ceftriaxone, cefepime, levofloxacin, amikacin and ertapenem minimum inhibitory concentrations (mg/L) distribution from community- and hospital-settings. Figure 1. Enterobacterales ceftriaxone and levofloxacin minimum inhibitory concentrations (mg/L) distribution from community- and hospital-settings. Conclusion Similar antimicrobials resistances were found in Enterobacterales from community- and hospital-acquired infections. New anti-infective agents are needed urgently to treat pathogens from the community-acquired infections and hospitals that have resistance to the first line regimen. Additionally, community antimicrobial stewardship programs are required. Disclosures All Authors: No reported disclosures

Sulfonamide-salicylaldehyde imines active against methicillin- and trimethoprim/sulfonamide-resistant Staphylococci

Background: Increasing resistance has resulted in an urgent need for new antimicrobial drugs. A systematic me-too approach was chosen to modify clinically used sulfonamides to obtain their imines. Methods & results: Twenty-five compounds were synthesized and evaluated for their antibacterial activity. The most active compounds were also investigated against methicillin- and trimethoprim/sulfamethoxazole (SMX)-resistant Gram-positive species. Staphylococci shared the highest susceptibility including resistant strains with minimum inhibitory concentrations from 3.91 μM (≥2.39 μg ml-1). Crucially, the compounds inhibit MRSA and trimethoprim/SMX-resistant Staphylococci without any cross-resistance. Modification of parent sulfonamides turned a bacteriostatic effect into a bactericidal effect. Toxicity for HepG2 and hemolytic properties were also determined. Conclusions: The presence of a dihalogenated salicylidene moiety is required for optimal activity. Based on toxicity, promising derivatives for further investigation were identified.

Antimicrobial resistance of Staphylococcus aureus strains isolated from cow raw milk samples from Albania and Serbia

This research aimed to determine the prevalence rate of antimicrobial resistance (AMR) of Staphylococcus aureus isolated from raw cow milk samples in Albania and Serbia. A total of 100 raw milk samples, 50 from Albania and 50 from Serbia, have been collected from randomly selected cattle farms. Twelve samples (12 %) were positive for S. aureus, five of those originating from Albania (41.66 %) and seven from Serbia (58.33 %). Resistance of isolated S. aureus strains to ampicillin in milk samples from Albania (36.7 %) and Serbia (34.1 %) was the most common. Some of S. aureus strains from milk samples from Albania were resistant to tetracycline (16.9 %), while isolates from milk samples from Serbia were more resistant towards oxacillin (18 %). The recorded resistance towards erythromycin (13.2 %; 13.1 %), and sulfatrim (7.6 %; 6.9 %) was similar between both milk samples, respectively. The obtained results have shown higher resistance of S. aureus strains towards cephalothin in milk samples from Serbia (3.9 %) compared to milk samples from Albania (1.6 %), respectively. All isolates tested for antibiotic sensitivity were susceptible to methicillin, vancomycin, chloramphenicol, and ciprofloxacin. The obtained results regarding the minimum inhibitory concentrations (MICs) of antibiotics indicated that isolates were resistant to tetracycline, oxacillin, erythromycin, and ampicillin, respectively. Results of our investigation have shown that the found antibiotics residues are related to the usage of erythromycin (72.6 % and 73.9 %), followed by ampicillin (70.4 % and 71.2 %) while residues of ciprofloxacin in the analysed milk samples was not registered.

Minimum inhibitory concentrations of commonly used antibiotics against Helicobacter Pylori: A multicenter study in South China

Aim To determine the minimum inhibitory concentrations (MICs) of commonly used antibiotics against Helicobacter Pylori (H. pylori) in South China and compare their resistance rates by using EUCAST breakpoints and other breakpoints. Methods Patients who had not previously received H. pylori treatment in clinical centers in South China were enrolled in this study from 2017 to 2020. Gastric biopsies were obtained for H. pylori culture. The MICs of amoxicillin (AMX), clarithromycin (CLA), metronidazole (MTZ), levofloxacin (LEV), tetracycline (TET) and furazolidone (FZD) were tested by broth microdilution method and assessed by two different breakpoints. ATCC43504 standard strain served as a control. Results A total of 208 H. pylori strains were isolated from patients’ biopsy samples. The MICs of AMX, CLA, MTZ, LEV, TET and FZD for H. pylori were 0.0156-256mg/L (MIC50 0.125mg/L, MIC90 4mg/L), 0.0156- >256 mg/L (MIC50 0.0312mg/L, MIC90 64mg/L), 0.0156- >256mg/L (MIC50 8mg/L, MIC90 256mg/L), 0.0156-256mg/L (MIC50 0.25mg/L, MIC90 16mg/L), 0.0156-256mg/L (MIC50 0.0625mg/L, MIC90 4mg/L), and 0.0156- >256mg/L (MIC50 0.0312mg/L, MIC90 2mg/L), respectively. The MICs of AMX, CLA, MTZ, LEV, TET and FZD for ATCC43504 strain were 0.25mg/L, 0.0625mg/L, 64mg/L, 0.5mg/L, 1mg/L and 0.25mg/L, respectively. The resistance rate of FZD was 11.05%. The overall resistance rates according to EUCAST breakpoints and other breakpoints were 57.21% and 14.90% for AMX (p<0.001), 38.94% and 38.94% for CLA (p = 1), 39.42% and 50.96% for MTZ (p<0.001), 12.98% and 10.58% for TET (p = 0.025), 35.10% and 35.10% for LEV (p = 1), respectively. Conclusions Our results demonstrate that AMX, FZD, and TET, but not MTZ, CLR or LEV, showed good anti-H. pylori activity in vitro in South China. When different breakpoints were used, similar results were found with CLA, and LEV, but not with AMX, MTZ, or TET.