Antimicrobial resistance patterns among critical priority pathogens in an intensive care unit at a tertiary hospital in Egypt: a descriptive analysis comparing pre- and COVID-19 eras.

Background. The intensified global challenge of antimicrobial resistance, set against the backdrop of the COVID-19 pandemic, is a cause for major concern. Within healthcare settings, intensive care units are recognized as focal points for Gram-negative infections. The study pursued to assess the prevalence and antimicrobial resistance patterns of critical priority pathogens (Acinetobacter baumannii, Pseudomonas aeruginosa, Enterobacteriaceae, comprising Klebsiella pneumoniae and Escherichia coli) during both pre- and COVID-19 periods.Gap Statement. The decision to explore this topic stemmed from the urgent need to understand how the exceptional healthcare crisis of COVID-19 affected AMR patterns.Methods. This was an observational retrospective analysis of 1056 clinical specimens obtained from 950 patients who were admitted to the Medical Intensive Care Unit at Kasr Al-Aini Hospital, Cairo University, Egypt.Results. In the period before COVID-19, 342 pathogenic isolates (135 K. pneumoniae, 83 P. aeruginosa, 76 A. baumannii and 48 E. coli) were obtained from samples collected from 450 patients. Conversely, during the COVID-19 period, 714 isolates (237 K. pneumoniae, 205 A. baumannii, 199 P. aeruginosa and 73 E. coli) were collected from the same number of patients. In the course of the pandemic, there is a slight increase in A. baumannii and P. aeruginosa infections, whereas E. coli and K. pneumoniae exhibit a distinct trend with a noticeable reduction in infection rates during COVID-19. During the COVID-19 period, a noticeable rise in resistance rates was observed for all antibiotics utilized. The results from Fisher's exact test indicated a substantial increase in resistance towards certain antibiotics. Specifically, a significant rise in resistance was observed for E. coli to ciprofloxacin (P = 0.00), gentamicin and P. aeruginosa (P = 0.02), levofloxacin and A. baumannii (P = 0.01), piperacillin-tazobactam and A. baumannii (P = 0.04), and piperacillin-tazobactam and P. aeruginosa (P = 0.01).Conclusion. Our results display how the pandemic impacted bacterial infections and antibiotic resistance, indicating a general increase in resistance rates. These findings are crucial for guiding healthcare practices, emphasizing the need for continued surveillance and potentially checking antibiotic usage schemes.

Retained colistin susceptibility in clinical Acinetobacter baumannii isolates with multiple mutations in pmrCAB and lpxACD operons.

The progressive increase in the resistance rates to first- and second-line antibiotics has forced the reuse of colistin as last-line treatment for Acinetobacter baumannii infections, but the emergence of colistin-resistant strains is not uncommon. This has been long linked to acquired chromosomal mutations in the operons pmrCAB and lpxACD. Hence, such mutations are routinely screened in colistin-resistant strains by most studies. The current study was designed to explore the possible existence of pmrCAB and lpxACD mutations in colistin-susceptible isolates. For this purpose, the whole genome sequences of eighteen multi-/extensively drug resistant A. baumannii were generated by Illumina sequencing and screened for missense mutations of the operons pmrCAB and lpxACD. Most of the isolates belonged to global clones (GCs) including GC1 (n=2), GC2 (n=7), GC7 (n=2), GC9 (n=3), and GC11 (n=1). The minimum inhibitory concentrations (MICs) of colistin were determined by the broth microdilution assay. Seventeen isolates were fully susceptible to colistin with MICs ranging from (≤0.125 to 0.5 µg/ml). Interestingly, all colistin-susceptible isolates carried missense mutations in pmrCAB and lpxACD operons with reference to A. baumannii ATCC 19606. Overall, 34 mutations were found. Most substitutions were detected in pmrC (n=20) while no mutations were found in pmrA or lpxA. Notably, the mutation pattern of the two operons was almost conserved among the isolates that belonged to the same sequence type (ST) or GC. This was also confirmed by expanding the analysis to include A. baumannii genomes deposited in public databases. Here, we demonstrated the possible existence of missense mutations in pmrCAB and lpxACD operons in colistin-susceptible isolates, shedding light on the importance of interpreting mutations with reference to colistin-susceptible isolates of the same ST/GC to avoid the misleading impact of the ST/GC-related polymorphism. In turn, this may lead to misinterpretation of mutations and, hence, overlooking the real players in colistin resistance that are yet to be identified.

Genetic Configuration of Genomic Resistance Islands in Acinetobacter baumannii Clinical Isolates From Egypt.

In Acinetobacter baumannii (A. baumannii), a wide repertoire of resistance genes is often carried within genomic resistance islands (RIs), particularly in high-risk global clones (GCs). As the first in Egypt, the current study aimed at exploring the diversity and genetic configuration of RIs in the clinical isolates of A. baumannii. For this purpose, draft genomes of 18 isolates were generated by Illumina sequencing. Disk diffusion susceptibility profiling revealed multidrug resistance (MDR) and extensive drug resistance (XDR) phenotypes in 27.7 and 72.2%, respectively. The highest susceptibility was noted for tigecycline (100.0%) followed by colistin (94.4%), for which an MIC50 of 0.25 µg/ml was recorded by the broth microdilution assay. Sequence typing (ST) showed that the majority of the isolates belonged to high-risk global clones (GC1, GC2, and GC9). A novel Oxford sequence type (ST2329) that also formed a novel clonal complex was submitted to the PubMLST database. A novel bla ADC variant (bla ADC-258) was also identified in strain M18 (ST85Pas/1089Oxf). In addition to a wide array of resistance determinants, whole-genome sequencing (WGS) disclosed at least nine configurations of genomic RIs distributed over 16/18 isolates. GC2 isolates accumulated the largest number of RIs (three RIs/isolate) followed by those that belong to GC1 (two RIs/isolate). In addition to Tn6022 (44.4%), the comM gene was interrupted by AbaR4 (5.5%) and three variants of A. baumannii genomic resistance island 1(AbGRI)-type RIs (44.4%), including AbaR4b (16.6%) and two novel configurations of AbGRI1-like RIs (22.2%). Three of which (AbaR4, AbaR4b, and AbGRI1-like-2) carried bla OXA-23 within Tn2006. With less abundance (38.8%), IS26-bound RIs were detected exclusively in GC2 isolates. These included a short version of AbGRI2 (AbGRI2-15) carrying the genes bla TEM-1 and aphA1 and two variants of AbGRI3 RIs carrying up to seven resistance genes [mphE-msrE-armA-sul1-aadA1-catB8-aacA4]. Confined to GC1 (22.2%), sulfonamide resistance was acquired by an ISAba1 bracketed GIsul2 RI. An additional RI (RI-PER-7) was also identified on a plasmid carried by strain M03. Among others, RI-PER-7 carried the resistance genes armA and bla PER-7. Here, we provided a closer view of the diversity and genetic organization of RIs carried by a previously unexplored population of A. baumannii.

Tn7382, a novel composite transposon harboring blaNDM-1 and aphA6 in Acinetobacter baumannii.

OBJECTIVES: Co-transfer of carbapenem and amikacin resistance might contribute to the evolution of extensively drug resistant (XDR) Acinetobacter baumannii. The current study used bioinformatic tools and an in silico approach to investigate the potential mobility of a novel composite transposon co-harboring blaNDM-1 and aphA6. METHODS: The transposon, named here Tn7382, was recently identified in the chromosomes of two XDR A. baumannii isolates (M02 and M11) from Egypt. The draft genomes of M02 and M11 were generated by Illumina sequencing. Nucleotide homology of Tn7382 and flanking regions was analyzed using the Basic Local Alignment Search Tool. RESULTS: Tn7382 is derived from Tn125 and encompasses seven open reading frames [aphA6, ISAba125 transposase-coding gene, blaNDM-1, ble, iso, tat, cutA] enclosed by two direct copies of ISAba14. While described for the first time, Tn7382 was found in the chromosomes of five A. baumannii strains deposited in the NCBI database. Using the Artemis Comparison Tool, the potential mobility of Tn7382 was demonstrated in silico by comparative genomic analysis of two A. baumannii strains (TP1 and TP2) retrieved from the NCBI database. The transposon was acquired by TP2 at the same location as an ISAba14 element in the ancestral variant TP1 isolated from the same patient in the USA 11 days earlier. CONCLUSIONS: Here, we present the characteristics of Tn7382, a composite transposon flanked by ISAba14 and harboring the aphA6 and blaNDM-1 resistance genes. In silico analysis inferred the potential mobility of Tn7382, but experimental validation is still required.

Genomic Characterization of Extensively Drug-Resistant NDM-Producing Acinetobacter baumannii Clinical Isolates With the Emergence of Novel bla ADC-257.

Acinetobacter baumannii has become a major challenge to clinicians worldwide due to its high epidemic potential and acquisition of antimicrobial resistance. This work aimed at investigating antimicrobial resistance determinants and their context in four extensively drug-resistant (XDR) NDM-producing A. baumannii clinical isolates collected between July and October 2020 from Kasr Al-Ainy Hospital, Cairo, Egypt. A total of 20 A. baumannii were collected and screened for acquired carbapenemases (bla NDM, bla VIM and bla IMP) using PCR. Four NDM producer A. baumannii isolates were identified and selected for whole-genome sequencing, in silico multilocus sequence typing, and resistome analysis. Antimicrobial susceptibility profiles were determined using disk diffusion and broth microdilution tests. All bla NDM-positive A. baumannii isolates were XDR. Three isolates belonged to high-risk international clones (IC), namely, IC2 corresponding to ST570Pas/1701Oxf (M20) and IC9 corresponding to ST85Pas/ST1089Oxf (M02 and M11). For the first time, we report bla NDM-1 gene on the chromosome of an A. baumannii strain that belongs to sequence type ST164Pas/ST1418Oxf. Together with AphA6, bla NDM-1 was bracketed by two copies of ISAba14 in ST85Pas isolates possibly facilitating co-transfer of amikacin and carbapenem resistance. A novel bla ADC allele (bla ADC-257) with an upstream ISAba1 element was identified in M19 (ST/CC164Pas and ST1418Oxf/CC234Oxf). bla ADC genes harbored by M02 and M11 were uniquely interrupted by IS1008. Tn2006-associated bla OXA-23 was carried by M20. bla OXA-94 genes were preceded by ISAba1 element in M02 and M11. AbGRI3 was carried by M20 hosting the resistance genes aph(3`)-Ia, aac(6`)-Ib`, catB8, ant(3``)-Ia, sul1, armA, msr(E), and mph(E). Nonsynonymous mutations were identified in the quinolone resistance determining regions (gyrA and parC) of all isolates. Resistance to colistin in M19 was accompanied by missense mutations in lpxACD and pmrABC genes. The current study provided an insight into the genomic background of XDR phenotype in A. baumannii recovered from patients in Egypt. WGS revealed strong association between resistance genes and diverse mobile genetic elements with novel insertion sites and genetic organizations.

The TACTIC experience: establishing an international, interdisciplinary network to tackle antimicrobial resistance.

Antimicrobial resistance (AMR) is a major global health threat that requires an interdisciplinary international approach to address. In response to calls from policymakers and funders alike, a growing number of research networks on AMR have been created with this approach in mind. However, there are many challenges facing researchers in establishing such networks and research projects. In this article, we share our experience of establishing the network 'TACTIC: Tackling AMR Challenges through Translational Interdisciplinary Collaborations'. Although presented with many challenges both scientific and logistical, the network has underpinned productive interaction between biomedical and social scientists from several countries and fostered true collaboration in an educative, stimulating and sustainable way that forms a platform for important research on AMR.

Identification of entry inhibitors with 4-aminopiperidine scaffold targeting group 1 influenza A virus.

Influenza A viruses (IAVs) cause seasonal flu and occasionally pandemics. The current therapeutics against IAVs target two viral proteins - neuraminidase (NA) and M2 ion-channel protein. However, M2 ion channel inhibitors (amantadine and rimantadine) are no longer recommended by CDC for use due to the emergence of high level of antiviral resistance among the circulating influenza viruses, and resistant strains to NA inhibitors (oseltamivir and zanamivir) have also been reported. Therefore, development of novel anti-influenza therapies is urgently needed. As one of the viral surface glycoproteins, hemagglutinin (HA) mediates critical virus entry steps including virus binding to host cells and virus-host membrane fusion, which makes it a potential target for anti-influenza drug development. In this study, we report the identification of compound CBS1116 with a 4-aminopiperidine scaffold from a chemical library screen as an entry inhibitor specifically targeting two group 1 influenza A viruses, A/Puerto Rico/8/34 (H1N1) and recombinant low pathogenic avian H5N1 virus (A/Vietnam/1203/04, VN04Low). Mechanism of action studies show that CBS1116 interferes with the HA-mediated fusion process. Further structure activity relationship study generated a more potent compound CBS1117 which has a 50% inhibitory concentration of 70 nM and a selectivity index of ~4000 against A/Puerto Rico/8/34 (H1N1) infection in human lung epithelial cell line (A549).