Carbapenem-resistant Acinetobacter Baumannii Isolates Research Articles

Molecular Identification of OXA Carbapenemase-Encoding Genes in Acinetobacter baumannii Isolated from Patients in Critical Care in Egypt.

Background: The emergence of carbapenem-resistant Acinetobacter baumannii (CRAB) in hospitals, particularly within critical care units, has garnered substantial global concern. CRAB commonly arises from the degradation by various ß-lactamases. Objective: We aimed to assess OXA-type carbapenemases in clinical isolates of A. baumannii obtained from an Egyptian tertiary care facility. Patients and Methods: This study examined 25 distinct A. baumannii strains collected from various clinical samples of patients in intensive care unit. Bacterial identification was conducted utilizing both traditional methods and the Vitek2 system. Antibiotic resistance profiles were assessed according to the European Committee on Antimicrobial Susceptibility Testing standards using the Vitek2 Compact automated system. Additionally, multiplex real-time polymerase chain reaction was used to identify the presence of blaOXA23, blaOXA24, blaOXA51, and blaOXA58 carbapenemase genes. Colistin susceptibility was assessed utilizing the broth microdilution method. Results: Carbapenem resistance was identified in 100% of the studied isolates. The blaOXA51 gene was detected in all A. baumannii strains. The gene blaOXA23 was identified in 22 strains (88%), whereas blaOXA24 and blaOXA58 were present in 15 strains (60%). All isolates, except one, co-harbored two or more OXA encoding genes. Colistin resistance was detected in 4 of 25 strains (16%). Conclusion: Our findings demonstrate the widespread distribution of CRAB isolates that co-harbor multiple carbapenemase-encoding genes. Molecular epidemiological studies and the surveillance of antibiotic resistance profiles may aid in identifying and tracing the origins of resistant bacteria, thereby limiting their spread.

Incidence of colistin heteroresistance among carbapenem-resistant Acinetobacter baumannii clinical isolates in a tertiary care hospital in Pakistan.

The emergence of colistin-resistant and heteroresistant strains of carbapenem-resistant Acinetobacter baumannii (CRAB) complicates treatment and exacerbates the global health crisis of drug-resistant bacteria. This study aims to investigate the incidence and clinical implications of colistin heteroresistance in carbapenem-resistant Acinetobacter baumannii isolates from a tertiary hospital in Pakistan. A total of 130 CRAB isolates were collected from December 2022 to December 2023. Colistin susceptibility was assessed using broth microdilution, and heteroresistance was detected through population analysis profiling. Heteroresistance (HR) was identified in 31.5% (41/130) of the isolates, while 7.7% were colistin-resistant, despite initial susceptibility indicated by broth microdilution. Clinical data revealed that HR was associated with significant 14-day clinical failure but not with 30-day all-cause mortality. Heteroresistant strains showed extensive multidrug resistance, posing a serious threat to effective treatment. The study highlights the critical need for accurate detection of colistin HR to prevent treatment failure and improve patient outcomes. The prevalence of colistin HR underscores the necessity for revised diagnostic and treatment strategies in Pakistan, emphasizing the importance of recognizing and addressing this emerging threat in healthcare settings.

A single-center analysis of clonal transmission of carbapenem-resistant Acinetobacter baumannii among intensive care unit patients during the COVID-19 pandemic

Carbapenem-Resistant Acinetobacter baumannii (CRAB) outbreak in intensive care units (ICUs) is a significant problem for healthcare facilities. In this study, we aimed to investigate the occurrence of CRAB isolates among ICU-admitted patients during the three waves of the COVID-19 pandemic in Iran using Multiple-Locus Variable Number Tandem-Repeat Analysis (MLVA). We obtained 50 (A) baumannii isolates from tracheal aspirate and blood culture samples. In the disc diffusion method, all isolates were cefotaxime, ceftriaxone, and cefepime-resistant, while 98% (49/50) of isolates were resistant to piperacillin, piperacillin-tazobactam, ceftazidime, and ciprofloxacin. Levofloxacin and tobramycin resistance was found in 76% (38/50) of isolates. In the microbroth dilution test all isolates were resistant to imipenem, 98% (49/50) to meropenem, 68% (34/50) to colistin, and 20% (10/50) to polymyxin (B) Based on the PCR findings, all isolates harbored blaOXA−40, ISAba-1, and int-2 genes. There were no isolates found that have the blaOXA−58, blaOXA−143, blaVEB−1, blaVIM, and int-3 genes. Among Extended-spectrum beta-lactamases (ESBL) genes, blaCTX−M, blaTEM, blaSHV, blaGES, and blaPER−1 have a prevalence of 42% (21/50), 84% (42/50), 58% (29/50), 78% (39/50), and 54% (27/50), respectively. 74% (37/50) of the isolates had the blaOXA−23 gene, while all of the isolates carried the blaOXA−40 gene. Among MBL genes, blaIPM, blaGIM, blaSIM, and blaNDM−1 have a prevalence of 20% (10/50), 8% (4/50), 22% (11/50), and 60% (30/50), respectively. The prevalence of int-1 was documented as 74% (37/50). Accordingly, all isolates were identified as CRAB. The co-existence of blaOXA−23/int-2 and blaOXA−23/isaba-1 was 74% (37/50). The co-existence of blaNDM−1/ISAba-1 was observed in 30 (60%) isolates. Using an 80% similarity threshold on the dendrogram constructed through MLVA typing, all isolates were grouped into two clusters: cluster A with 9 isolates from wave 3, and cluster B with 41 isolates from waves 3, 4, and 5. Our study confirms a clonal transmission of CRAB during the study period and suggests using molecular typing methods like MLVA in healthcare settings to identify dominant clones, antibiotic resistance patterns, and transmission routes. This will help to better manage the emergence and spread of antibiotic-resistant strains in future outbreaks.

Phenotypic and molecular characterization of carbapenem-resistant Acinetobacter baumannii isolates from ventilator associated pneumonia patients

Phenotypic and molecular characterization of carbapenem-resistant Acinetobacter baumannii isolates from ventilator associated pneumonia patients

Handwashing sinks as reservoirs of carbapenem-resistant Acinetobacter baumannii in the intensive care unit: a prospective multicenter study.

The extent to which sinks are contaminated by carbapenem-resistant Acinetobacter baumannii (CRAB) in intensive care units (ICUs) and the association between these contaminated sinks and hospital-acquired CRAB infections during the non-cluster period remains largely unknown. Here, we performed a prospective multicenter study in 16 ICUs at 11 tertiary hospitals in Chengdu, China. We sampled sinks, collected CRAB clinical isolates, and conducted whole-genome sequencing and analysis. A total of 789 swabs were collected from 158 sinks, and 16 CRAB isolates were recovered from 16 sinks, resulting in a contamination rate of 10.16%. Twenty-seven clinical isolates were collected during the study period. The majority (97.67%, 42/43) of the CRAB isolates belonged to ST2, and 36 (83.72%) of them had both bla OXA-23 and bla OXA-66. The 43 strains belonged to 12 clones. One certain clone caused multiple contaminations of seven sinks in one GICU. Two clones of ST2 bla OXA-23 and bla OXA-66-carrying sink strains were likely the sources of the two clusters in the two GICUs, respectively. Five ST2 bla OXA-23-carrying isolates were found to be common clones but were recovered from two hospitals. The contamination rate of CRAB in handwashing sinks is high in some local ICUs, and the contaminated sinks can serve as environmental reservoirs for CRAB clusters.

Molecular Detection of Carbapenemases in Acinetobacter baumannii Strains of Portugal and Association With Sequence Types, Capsular Types, and Virulence

Carbapenem-resistant Acinetobacter baumannii (CRAB) is an important nosocomial pathogen. The capsular type (K-type) is considered a major virulence factor, contributing to the evasion of host defenses. The global spread and dissemination dynamics between K-types, sequence types (ST), antibiotic resistance genes, and virulence factors remain largely unknown in Portugal. A collection of 96 CRAB clinical samples collected between 2005 and 2019 in the northern region of Portugal were tested for antimicrobial susceptibility profile and screened by polymerase chain reaction for resistance genetic determinants. A subset of 26 representative isolates was subjected to whole-genome sequencing to assess K types, ST types, and genomic relatedness. The pathogenicity of distinct K-types was also tested using Galleria mellonella model. For the 96 CRAB isolates analyzed, high antimicrobial resistance (>90%) was observed to the carbapenems, fluoroquinolones, and miscellaneous agents. Greater antimicrobial susceptibility (∼30%-57%) was observed for aminoglycosides, particularly tobramycin, and amikacin. Genotypically, 75 strains (78.5%) carried blaOXA-23-like, 18 strains (18.8%) carried blaIMP-like, and 11 strains (14.9%) carried blaOXA-40-like carbapenem resistance genes, respectively. Associations between OXA and ST/capsular locus (KL) types were observed over the years (eg, OXA-40-like/ST46Past/KL120 and OXA-23-like/ST2Past/KL2). ST2Past of clonal complex II was present in most strains, a dominant drug-resistant lineage in the United States and Europe. KL7 was also the most prevalent KL-type (38.5%), followed by KL2 (34.6%), KL120 (23.1%), and KL9 (3.8%). Virulence assessment for different K-types in a Galleria mellonella model revealed a significantly increased virulence for KL120 when compared with KL7, KL9, and KL2. There are specific CRAB serotypes circulating in Portugal, accounting by the low diversity of acquired carbapenemase genes (OXA-23-like and OXA-40-like), ST types (ST2 and ST46) and KL types (KL2, KL7, KL9, and KL120) identified. The high prevalent of ST2, especially when associated with KL2 and blaOXA-23-like, suggest that antibiotic resistance has been driven by clonal expansion of clonal complex II. Such findings provide useful information on the diversity of multidrug-resistant bacterium that might be relevant for antibacterial interventions.

Comparison of antimicrobial activities and resistance mechanisms of eravacycline and tigecycline against clinical Acinetobacter baumannii isolates in China.

Tigecycline (TGC) is currently used to treat carbapenem-resistant Acinetobacter baumannii (CRAB) infections, while eravacycline (ERV), a new-generation tetracycline, holds promise as a novel therapeutic option for these infections. However, differences in resistance mechanism between ERV and TGC against A. baumannii remain unclear. This study sought to compare the characteristics and mechanisms of ERV and TGC resistance among clinical A. baumannii isolates. A total of 492 isolates, including 253 CRAB and 239 carbapenem-sensitive A. baumannii (CSAB) isolates, were collected from hospitalized patients in China. The MICs of ERV and TGC against A. baumannii were determined by broth microdilution. Genetic mutations and expressions of adeB, adeG, adeJ, adeS, adeL, and adeN in resistant strains were examined by PCR and qPCR, respectively. The in vitro recombination experiments were used to verify the resistance mechanism of ERV and TGC in A. baumannii. The MIC90 of ERV in CRAB and CSAB isolates were lower than those of TGC. A total of 24 strains resistant to ERV and/or TGC were categorized into three groups: only ERV-resistant (n = 2), both ERV- and TGC-resistant (n = 7), and only TGC-resistant (n = 15). ST208 (75%, n = 18) was a major clone that has disseminated in all three groups. The ISAba1 insertion in adeS was identified in 66.7% (6/9) of strains in the only ERV-resistant and both ERV- and TGC-resistant groups, while the ISAba1 insertion in adeN was found in 53.3% (8/15) of strains in the only TGC-resistant group. The adeABC and adeRS expressions were significantly increased in the only ERV-resistant and both ERV- and TGC-resistant groups, while the adeABC and adeIJK expressions were significantly increased and adeN was significantly decreased in the only TGC-resistant group. Expression of adeS with the ISAba1 insertion in ERV- and TGC-sensitive strains significantly increased the ERV and TGC MICs and upregulated adeABC and adeRS expressions. Complementation of the wildtype adeN in TGC-resistant strains with the ISAba1 insertion in adeN restored TGC sensitivity and significantly downregulated adeIJK expression. In conclusion, our data illustrates that ERV is more effective against A. baumannii clinical isolates than TGC. ERV resistance is correlated with the ISAba1 insertion in adeS, while TGC resistance is associated with the ISAba1 insertion in adeN or adeS in A. baumannii.

Characterization of the carbapenem-resistant Acinetobacter baumannii clinical reference isolate BAL062 (CC2:KL58:OCL1): resistance properties and capsular polysaccharide structure.

The carbapenem-resistant Acinetobacter baumannii isolate BAL062 is a clinical reference isolate used in several recent experimental studies. It is from a ventilator-associated pneumonia (VAP) patient in an intensive care unit at the Hospital for Tropical Diseases (HTD), Ho Chi Minh City, Vietnam in 2009. Here, BAL062 was found to belong to the B sub-lineage of global clone 2 (GC2) isolates in the previously reported outbreak (2008 and 2012) of carbapenem-resistant VAP A. baumannii at the HTD. While related sub-lineage B outbreak isolates were extensively antibiotic-resistant and carry GC2-associated genomic resistance islands, AbGRI1, AbGRI2, and AbGRI3, BAL062 has lost AbGRI3 and three aminoglycoside resistance genes, armA, aacA4, and aphA1, leading to amikacin, tobramycin and kanamycin susceptibility. The location of Tn2008VAR found in the chromosome of this sub-lineage was also corrected. Like many of the outbreak isolates, BAL062 carries the KL58 gene cluster at the capsular polysaccharide (CPS) synthesis locus and an annotation key is provided. As information about K type is important for the development of novel CPS-targeting therapies, the BAL062 K58-type CPS structure was established using NMR spectroscopy. It is most closely related to K2 and K93, sharing similar configurations and linkages between K units, and contains the rare higher monosaccharide, 5,7-diacetamido-3,5,7,9-tetradeoxy-d-glycero-l-manno-non-2-ulosonic acid (5,7-di-N-acetyl-8-epipseudaminic acid; 8ePse5Ac7Ac), the 8-epimer of Pse5Ac7Ac (5,7-di-N-acetylpseudaminic acid). Inspection of publicly available A. baumannii genomes revealed a wide distribution of the KL58 locus in geographically diverse isolates belonging to several sequence types that were recovered over two decades from clinical, animal, and environmental sources.IMPORTANCEMany published experimental studies aimed at developing a clearer understanding of the pathogenicity of carbapenem-resistant Acinetobacter baumannii strains currently causing treatment failure due to extensive antibiotic resistance are undertaken using historic, laboratory-adapted isolates. However, it is ideal if not imperative that recent clinical isolates are used in such studies. The clinical reference isolate characterized here belongs to the dominant A. baumannii GC2 clone causing extensively resistant infections and has been used in various recent studies. The correlation of resistance profiles and resistance gene data is key to identifying genes available for gene knockout and complementation analyses, and we have mapped the antibiotic resistance genes to find candidates. Novel therapies, such as bacteriophage or monoclonal antibody therapies, currently under investigation as alternatives or adjuncts to antibiotic treatment to combat difficult-to-treat CRAb infections often exhibit specificity for specific structural epitopes of the capsular polysaccharide (CPS), the outer-most polysaccharide layer. Here, we have solved the structure of the CPS type found in BAL062 and other extensively resistant isolates. As consistent gene naming and annotation are important for locus identification and interpretation of experimental studies, we also have correlated automatic annotations to the standard gene names.

Frequency of cefiderocol heteroresistance among patients treated with cefiderocol for carbapenem-resistant Acinetobacter baumannii infections.

Cefiderocol exhibits potent in vitro activity against carbapenem-resistant Acinetobacter baumannii (CRAb), but this activity has not consistently translated to improved outcomes among patients. Cefiderocol heteroresistance, or the presence of a resistant subpopulation, has been proposed as one possible explanation. The objective of this study was to explore associations between heteroresistance and outcomes of patients with CRAb infections. Baseline CRAb isolates were collected from 27 consecutive patients in the USA and Italy. Cefiderocol susceptibility was tested by broth microdilutions in triplicate. Heteroresistance was defined by population analysis profiling in duplicate. Resistance mechanisms and strain relatedness were evaluated through comparative genomic analysis. Overall, 59% of infecting CRAb isolates were identified as cefiderocol-heteroresistant; rates were higher among isolates from Italy (79%) than the USA (38%). The median Charlson Comorbidity and SOFA scores were 4 and 5, respectively; 44% of patients had pneumonia, which was the most common infection type. Rates of 28-day clinical success and survival were 30% and 73%, respectively. By broth microdilution, cefiderocol MICs ≥1 mg/L were associated with higher failure rates than MICs ≤0.5 mg/L (81% versus 55%). Rates of clinical failure were numerically higher among patients infected by cefiderocol-heteroresistant compared with susceptible CRAb (81% versus 55%). Whole-genome sequencing identified a premature stop codon in the TonB-dependent receptor gene piuA in six isolates, all of which were heteroresistant. This pilot study supports the hypothesis that cefiderocol treatment failure may be associated with higher MICs and/or the presence of heteroresistance. Further studies are needed to confirm these findings.

Sequential use of capsular typing and whole-genome sequencing-based analysis for transmission of carbapenem-resistant Acinetobacter baumannii in a tertiary medical center

BackgroundDuring the COVID-19 pandemic, there has been an increasing trend in healthcare-associated infections (HAIs) caused by carbapenem-resistant Acinetobacter baumannii (CRAB), posting a global public health concern. The heightened sensitivity of whole-genome sequencing (WGS) renders it an optimal and potent tool for monitoring outbreaks and tracing the transmission routes of nosocomial pathogens. MethodWe collected CRAB isolates from March 1, 2023, to April 6, 2023 in Chang Gung Memorial Hospital Lin Kou branch, a tertiary medical center in northern Taiwan. Any two or more isolates with the same identifiable capsular K-locus (KL) types were selected, and analyzed via WGS to identify putative transmission clusters, combined with epidemiologic and retrospective analysis on medical records to confirm risk factors and hidden transmission chains. ResultA total of 48 non-redundant CRAB isolates were collected, belonging to ST2 of Pasteur MLST scheme and identifiable KL types of KL2, KL3, KL9, KL10, KL22, KL52. Excluding the KL types that was only found in 1 case, KL2 (n = 9, 22.5 %), KL3 (n = 24, 60 %), KL9 (n = 3, 7.5 %), and KL10 (n = 4, 10 %) were selected for further WGS analysis. Four distinct transmission clusters comprised of 2, 3, 10, and 23 cases were identified on a basis of phylogenetic status. 12 probable transmission chains were revealed, and 2 hidden transmission routes can be speculated. ConclusionThis study referred to some hidden transmission chains that may be missed from traditional surveillance measures. Despite its low prevalence and high cost currently, implementing WGS could be a efficient, prompt, and unequivocal option for future MDRO infection control.

Challenges Facing Two Outbreaks of Carbapenem-Resistant Acinetobacter baumannii: From Cefiderocol Susceptibility Testing to the Emergence of Cefiderocol-Resistant Mutants.

Carbapenem-resistant Acinetobacter baumannii (CRAB) infections are associated with poor outcomes depending on patient's conditions, clinical severity and type of infection, and treatment is challenging given the limited therapeutic options available. The aim of this study was to describe the clinical and microbiological characteristics of two outbreaks caused by CRAB in an intensive care unit (ICU). In addition, the mechanisms of resistance detected in these strains and the treatment chosen according to the available therapeutic options were analyzed. Overall, 28 patients were included. Ten patients (35.71%) had ventilator-associated pneumonia (VAP), ten (35.71%) had a bloodstream infection (BSI), and eight (28.57%) were only colonized. Recurrent infection occurred in 25% (5/20) of infected patients. Two different strains of A. baumannii were isolated from the index patient of the first outbreak. The first strain belonged to the ST85 and carried the blaNDM-1 carbapenemase gene, while the second belonged to the ST2 and carried blaOXA-23, and blaOXA-66 carbapenemase genes. The phylogenetic analysis revealed that the ST2 strain was the cause of the major outbreak, and mutations in the AmpC gene were related to progressive increasing minimum inhibitory concentration (MIC) and finally, cefiderocol-resistance in one strain. The CRAB isolates from the second outbreak were also identified as ST2. Cefiderocol-resistant strains tests identified by the disc diffusion method were involved in 24% (6/25) of nosocomial infections. Using broth microdilution (BMD) ComASP® only, 33.3% (2/6) of these strains were cefiderocol-resistant. All-cause ICU mortality was 21.4%. Conclusions: Cefiderocol is the first approved siderophore cephalosporin for the treatment of CRAB infections. Cefiderocol-resistant strains were related with blaNDM-1 carbapenemase and mutations in the AmpC gene. Cefiderocol-resistant strains or that cannot be properly interpreted by disk diffusion, should be retested using BMD for definitive categorization.

Molecular epidemiology of Acinetobacter baumannii during COVID-19 at a hospital in northern China

BackgroundThe wide spread of carbapenem-resistance clones of Acinetobacter baumannii has made it a global public problem. Some studies have shown that the prevalence of Acinetobacter baumannii clones can change over time. However, few studies with respect to the change of epidemiological clones in Acinetobacter baumannii during Corona Virus Disease 2019 (COVID-19) were reported. This study aims to investigate the molecular epidemiology and resistance mechanisms of Acinetobacter baumannii during COVID-19.ResultsA total of 95 non-replicated Acinetobacter baumannii isolates were enrolled in this study, of which 60.0% (n = 57) were identified as carbapenem-resistant Acinetobacter baumannii (CRAB). The positive rate of the blaOXA−23 gene in CRAB isolates was 100%. A total of 28 Oxford sequence types (STs) were identified, of which the most prevalent STs were ST540 (n = 13, 13.7%), ST469 (n = 13, 13.7%), ST373 (n = 8, 8.4%), ST938 (n = 7, 7.4%) and ST208 (n = 6, 6.3%). Differently, the most widespread clone of Acinetobacter baumannii in China during COVID-19 was ST208 (22.1%). Further study of multidrug-resistant ST540 showed that all of them were carrying blaOXA−23, blaOXA−66, blaADC−25 and blaTEM−1D, simultaneously, and first detected Tn2009 in ST540. The blaOXA−23 gene was located on transposons Tn2006 or Tn2009. In addition, the ST540 strain also contains a drug-resistant plasmid with msr(E), armA, sul1 and mph(E) genes.ConclusionThe prevalent clones of Acinetobacter baumannii in our organization have changed during COVID-19, which was different from that of China. ST540 strains which carried multiple drug-resistant mobile elements was spreading, indicating that it is essential to strengthen the molecular epidemiology of Acinetobacter baumannii.

Nosocomial transmission of tet(x3), bla NDM-1 and bla OXA-97-carrying Acinetobacter baumannii conferring resistance to eravacycline and omadacycline, the Netherlands, March to August 2021.

Carbapenem-resistant Acinetobacter baumannii (CRAb) is an important pathogen causing serious nosocomial infections. We describe an outbreak of CRAb in an intensive care unit in the Netherlands in 2021. During an outbreak of non-resistant A. baumannii, while infection control measures were in place, CRAb isolates carrying highly similar bla NDM-1 - and tet(x3)-encoding plasmids were isolated from three patients over a period of several months. The chromosomal and plasmid sequences of the CRAb and non-carbapenemase-carrying A. baumannii isolates cultured from patient materials were analysed using hybrid assemblies of short-read and long-read sequences. The CRAb isolates revealed that the CRAb outbreak consisted of two different strains, carrying similar plasmids. The plasmids contained multiple antibiotic resistance genes including the tetracycline resistance gene tet(x3), and the bla NDM-1 and bla OXA-97 carbapenemase genes. We determined minimal inhibitory concentrations (MICs) for 13 antibiotics, including the newly registered tetracycline antibiotics eravacycline and omadacycline. The CRAb isolates showed high MICs for tetracycline antibiotics including eravacycline and omadacycline, except for minocycline which had a low MIC. In this study we show the value of sequencing multidrug-resistant A. baumannii for outbreak tracking and guiding outbreak mitigation measures.

Survey of VA Laboratory Practices for Carbapenem-resistant Acinetobacter baumannii and Pseudomonas aeruginosa

Background: Carbapenem-resistant Acinetobacter baumannii (CRAB) and Pseudomonas aeruginosa (CRPA) are drug-resistant pathogens causing high mortality rates with limited treatment options. Understanding the incidence of these organisms and laboratory knowledge of testing protocols is important for controlling their spread in healthcare settings. This project assessed how often Veterans Affairs (VA) healthcare facilities identify CRAB and CRPA and testing practices used. Method: An electronic survey was distributed to 126 VA acute care facilities September-October 2023. The survey focused on CRAB and CRPA incidence, testing and identification, and availability of testing resources. Responses were analyzed by complexity of patients treated at VA facilities (High, Medium, Low) using Fisher’s exact tests. Result: 77 (61.1%) facilities responded, most in urban settings (85.4%). Most respondents were lead or supervisory laboratory technologists (84.2%) from high complexity facilities (69.0%). Few facilities detected CRAB ≥ once/month (4.4%), with most reporting that they have not seen CRAB at their facility (55.0%). CRPA was detected more frequently: 19% of facilities with isolates ≥ once/month, 29.2% a few times per year, and 26.9% reporting had not seen the organism. No differences in CRAB or CRPA incidence was found by facility complexity. Nearly all facilities, regardless of complexity, utilize the recommended methods of MIC or disk diffusion to identify CRAB or CRPA (91.9%) with remaining facilities reporting that testing is done off-site (7.8%). More high complexity facilities perform on-site testing compared to low complexity facilities (32.0% vs 2.7%, p=0.04). 83% of laboratories test for Carbapenemase production, with one-fourth using off-site reference labs. One-fourth of facilities perform additional antibiotic susceptibility testing for CRAB and CRPA isolates, most of which test for susceptibility to combination antibiotics; no differences between complexities were found. Agreement that sufficient laboratory and equipment resources were available was higher in high complexity than in medium complexity facilities (70.7% vs 33.3%, p=0.01), but not low complexity facilities (43.8%). Conclusion: Having timely and accurate testing protocols for CRAB and CRPA are important to quickly control spread and reduce associated mortality. This study shows that most VA protocols follow recommended testing and identification guidelines. Interestingly, there was no difference in CRAB or CRPA incidence for facilities providing higher vs lower complexity of care. While high and low complexity facilities generally reported sufficient resources for CRAB and CRPA evaluation, some medium-complexity labs, who may feel more compelled than low-complexity labs to bring testing in house, reported that additional resources would be required.

Isolation and Characterization of Novel Bacteriophages to Target Carbapenem-Resistant Acinetobacter baumannii.

The spread of multidrug-resistant Acinetobacter baumannii in hospitals and nursing homes poses serious healthcare challenges. Therefore, we aimed to isolate and characterize lytic bacteriophages targeting carbapenem-resistant Acinetobacter baumannii (CRAB). Of the 21 isolated A. baumannii phages, 11 exhibited potent lytic activities against clinical isolates of CRAB. Based on host spectrum and RAPD-PCR results, 11 phages were categorized into four groups. Three phages (vB_AbaP_W8, vB_AbaSi_W9, and vB_AbaSt_W16) were further characterized owing to their antibacterial efficacy, morphology, and whole-genome sequence and were found to lyse 37.93%, 89.66%, and 37.93%, respectively, of the 29 tested CRAB isolates. The lytic spectrum of phages varied depending on the multilocus sequence type (MLST) of the CRAB isolates. The three phages contained linear double-stranded DNA genomes, with sizes of 41,326-166,741 bp and GC contents of 34.4-35.6%. Genome-wide phylogenetic analysis and single gene-based tree construction revealed no correlation among the three phages. Moreover, no genes were associated with lysogeny, antibiotic resistance, or bacterial toxins. Therefore, the three novel phages represent potential candidates for phage therapy against CRAB infections.

Can flow cytometric measurements of reactive oxygen species levels determine minimal inhibitory concentrations and antibiotic susceptibility testing for Acinetobacter baumannii?

Current antimicrobial susceptibility testing (AST) requires 16-24 hours, delaying initiation of appropriate antibiotics. Hence, there is a need for rapid AST. This study aims to develop and evaluate the feasibility of a rapid flow cytometric AST assay to determine minimum inhibitory concentration (MIC) for carbapenem-resistant Acinetobacter baumannii (CRAB). Antibiotic exposure causes increased intracellular reactive oxygen species (ROS) in bacteria. We hypothesized that ROS can be used as a marker to determine MIC. We assessed three CRAB clinical isolates across fifteen antibiotics at various concentrations in a customized 96-well microtiter plate. The antibiotics assessed include amikacin, beta-lactams (ampicillin/sulbactam, aztreonam, cefepime, ceftolozane/tazobactam, doripenem, imipenem, meropenem, and piperacillin/tazobactam), levofloxacin, polymyxin B, rifampicin, trimethoprim/sulfamethoxazole, and tetracyclines (tigecycline and minocycline). These clinical CRAB isolates were assessed for ROS after antibiotic treatment. Increased ROS levels indicated by increased RedoxSensorTM Green (RSG) fluorescence intensity was assessed using flow cytometry (FCM). MIC was set as the lowest antibiotic concentration that gives a ≥1.5-fold increase in mode RSG fluorescence intensity (MICRSG). Accuracy of MICRSG was determined by comparing against microtiter broth dilution method performed under CLSI guidelines. ROS was deemed accurate in determining the MICs for β-lactams (83.3% accuracy) and trimethoprim/sulfamethoxazole (100% accuracy). In contrast, ROS is less accurate in determining MICs for levofloxacin (33.3% accuracy), rifampicin (0% accuracy), amikacin (33.3% accuracy), and tetracyclines (33.3% accuracy). Collectively, this study described an FCM-AST assay to determine antibiotic susceptibility of CRAB isolates within 5 hours, reducing turnaround time up to 19 hours.

Four genotypes of carbapenem-resistant Acinetobacter baumannii strains lacking OXA-23 production in Korea

During nationwide Fantimicrobial surveillance (Korea Global Antimicrobial Resistance Surveillance System [Kor-GLASS]), the recent emergence of non-oxacillinase (OXA)-23 production by carbapenem-resistant Acinetobacter baumannii (CRAB) isolates was noted. In this study, we evaluated resistance mechanisms other than OXA-23 production to elucidate the shift in considerable CRAB clones. The presence of OXA carbapenemase genes, such as blaOXA-23, blaOXA-24, blaOXA-58, and blaOXA-51-ISAba1, was determined by PCR. Other carbapenemase genes, such as blaIMP, blaVIM, blaNDM, blaKPC, blaGES, and blaOXA-48 , were determined using sequencing. Strains lacking carbapenemase genes were subjected to whole genome sequencing, and resistance genes were analyzed using ResFinder. Four CRAB strains were collected through a Kor-GLASS study in 2022, in which OXA-23 production was not identified. The carbapenemase genotypes of the four CRAB strains lacking blaOXA-23 were blaOXA-51-ISAba1, blaOXA-66/ACD25, blaOXA-182, and blaNDM-1. To the best of our knowledge, this is the f irst study to identify CRAB producing New Delhi metallo-β-lactamase (NDM)-1 in Korea. In conclusion, domestic CRAB resistance mechanisms may undergo subtle changes. Continuous observations are required to monitor the emergence of new clones.

Comparative study of phenotypic-based detection assays for carbapenemases in Acinetobacter baumannii

BackgroundAcinetobacter baumannii is a serious health concern worldwide, causing high mortality rates and limited medical therapy options. Carbapenem resistance is a significant problem in Acinetobacter baumannii isolates. The synthesis of acquired carbapenemases, such as oxacillinases, IMP, NDM, VIM, and KPC enzymes, causes carbapenem resistance. MethodsA total of 106 non-repetitive, Acinetobacter baumannii isolates were collected from four major hospitals in Bahrain including 78 carbapenem-resistant Acinetobacter baumannii (CRAB), and 28 carbapenem-susceptible Acinetobacter baumannii (CSAB) isolates. Three phenotypic tests were investigated in this study: including CARBA NP, modified carbapenem inactivation method (mCIM)/EDTA-CIM (eCIM), and modified Hodge test (MHT). ResultsCARBA NP was positive in 50 tested CRAB isolates (100%), and the sensitivity was 100%. The MHT was positive in 73/106 isolates (68.8%), while the sensitivity and specificity of the MHT were 77.6% and 100%. Moreover, only 38/106 (35.8%) isolates were positive for mCIM/eCIM. The sensitivity and specificity of mCIM were 40.4% and 100%. ConclusionCARBA NP was ideal for phenotypic detection of carbapenemase production, followed by MHT. The m/eCIM demonstrated a lower detection rate in CRAB. Consequently, combining tests would be more accurate. The mCIM/eCIM can easily distinguish between MBLs and serine-carbapenemases due to the frequent co-production of these enzymes in A. baumannii. In hospital setups where molecular characterization tests are not available, CARBA NP seems to be an alternative test in combination with MHT or mCIM/eCIM.

Multicenter retrospective genomic characterization of carbapenemase-producing Acinetobacter baumannii isolates from Jiangxi patients 2021-2022: identification of a novel international clone, IC11.

Carbapenem-resistant Acinetobacter baumannii (CRAB) is notorious for causing difficult-to-treat infections. To elucidate the molecular and clinical epidemiology of CRAB in Jiangxi, clinical CRAB isolates were collected and underwent whole-genome sequencing and antibiotic susceptibility phenotyping. Key findings included the predominance of OXA-23-producing IC2 A. baumannii, marked by the emergence of OXA-23 and NDM-1-producing IC11 strains.

Treatment-emergent cefiderocol resistance in carbapenem-resistant Acinetobacter baumannii is associated with insertion sequence ISAba36 in the siderophore receptor pirA.

We report the emergence of cefiderocol resistance in a blaOXA-72 carbapenem-resistant Acinetobacter baumannii isolate from a sacral decubitus ulcer. Cefiderocol was initially used; however, a newly approved sulbactam-durlobactam therapy with source control and flap coverage was successful in treating the infection. Laboratory investigation revealed cefiderocol resistance mediated by ISAba36 insertion into the siderophore receptor pirA.