Carbapenem resistant Acinetobacter baumannii (CRAB) is one of the major global threats for healthcare settings worldwide, as there are only few antibiotics effective to treat the infections caused by these isolates due to its high rates of antimicrobial resistance (Ejaz et al., 2021). This pathogen is responsible for multiple nosocomial infections such as bloodstream infections, urinary tract infections, wound infections, ventilator-associated pneumonia and other respiratory tract infections, meningitis and bacteremia (Kurihara et al., 2020). As a consequence, A. baumannii is heading the World Head Organization’s (WHO) list of critical pathogens for which new antibiotics are urgently needed (El-Kholy et al., 2021). Many other global institutions such as the European Centre for Disease Prevention and Control (ECDC), Infectious Diseases Society of America (IDSA) and the Center for Disease Control and Prevention of America (CDC) have also declared it an urgent threat (Mea et al., 2021).

Rates of mortality and disability caused by A. baumannii infections are increasing. Retrospective studies showed that the mortality rates associated with A. baumannii infections are ranging from 22.8% to 49.6% in the United States (US), and from 29% to 71.6% in Europe (Patel et al., 2019). According to autores, mortality associated with A. baumannii causing hospital-acquired and ventilator-associated pneumonia was higher in Western Asia (56.2%), Southern Europe (55.7%) and Northern Africa (53.3%). Countries of the Mediterranean area, such as Greece (68.2%), Turkey (61.4) or Egypt (53.3%) were the ones with the highest reported mortality rates (Mohd Sazlly Lim et al., 2019). Indeed, in Egypt 30–100% of A. baumannii isolates are considered as Multidrug-Resistant (MDR), and carbapenem resistance was reported in 26.6–100% of A. baumannii isolates (El-Kholy et al., 2021). Furthermore, many reports showed that the COVID-19 pandemic increased CRAB infection rates, for instance in the US, the rates in hospitals increased a 78% and overall by 35% in 2020 compared with 2019 (Centers for Disease Control and Prevention, 2022). In Egypt, MDR A. baumannii was the second most common cause of infection (27.4%) among mechanically ventilated patients during the second wave of COVID-19 (Elwakil et al., 2023). The economic impact is also remarkable in CRAB infections, for instance in the United States, the CDC reported that treatment costs were around $281 million in 2019 (Ejaz et al., 2021).

Different resistance mechanisms are commonly found in A. baumannii including antibiotic inactivation enzymes, alteration of target sites, overexpression of efflux pumps and loss of porins (Chakravarty, 2020). Carbapenemases are the main carbapenem resistance mechanism in A. baumannii, being carbapenem-hydrolyzing oxacilinases the most important ones. However, during the last years, class B metallo-β-lactamases such as New Delhi Metallo-Beta-lactamases (NDM) are on the rise (Xanthopoulou et al., 2020).

Nine major International Clones (IC1-9) of A. baumannii, have been described up to now, being IC2/CC92 with the acquired bla _OXA-23 gene the most disseminated lineage worldwide (Al-Hassan et al., 2019; Al-Hassan et al., 2021); and IC1 and IC2 in Europe (Muthuirulandi Sethuvel et al., 2019). In the Middle East and North Africa, A. baumannii clinical outbreaks, caused by MDR isolates endemically producing carbapenemases NDM-1, NDM-2 and OXA-23, are usually poly-clonal but with dominance of IC2 lineage (Hassan et al., 2021). Although IC2 is the most disseminated clone in Northern Africa, isolates belonging to IC5 and IC9 are also reported (Al-Hassan et al., 2021; Hamed et al., 2022). From 2013 to date, isolates belonging to Oxford ST208 (IC2), are commonly reported in Egyptian hospitals (Al-Hassan et al., 2019). However, in spite of the alarming situation, there is little information about A. baumannii in Egypt and more studies are needed (Hassan et al., 2021). Nevertheless, the limited resources for research in low- and middle-income countries such as Egypt, makes it difficult to obtain epidemiological data. It is of high concern to investigate the molecular epidemiology to control the dissemination of these clinically important isolates.

The aim of the present study was to characterize the genetic features, to study the molecular epidemiology and to identify the antimicrobial susceptibility profiles of thirty-six carbapenem resistant A. baumannii clinical isolates obtained from hospitals from Alexandria, Egypt.

It is said that the lack of regulation and the abuse in the use of antibiotics in Egypt are the main cause of the acceleration in the emergence of resistant isolates, and also responsible for exporting resistance to other countries (Elwakil et al., 2023). In the present study, 94.4% of carbapenem, 100% of fluoroquinolone and 86.11% of aminoglycoside resistance was observed, which is consistent with previous studies reporting a carbapenem resistance of 98% with elevated levels of resistance to quinolones and aminoglycosides in Mansoura, Egypt (Said et al., 2018). In fact, our isolates showed higher aminoglycoside resistance ratios than the resistance reported in recent studies from Egypt (up to 82% and 67%) (ELsheredy et al., 2021; Kishk et al., 2021). To our knowledge, up to date, no cefiderocol resistant isolates have been reported in Egypt apart from the eight resistant isolates analyzed in our study. Fortunately, isolates remained susceptible to colistin, although colistin resistance was reported in up to 53% of A. baumannii Egyptian isolates according to studies published in 2020 (Fam et al., 2020; Makharita et al., 2020).

WGS analysis showed bla _OXA-66 and bla _OXA-65 as the most frequent bla _OXA-51-like variants among the isolates, which is consistent with the high prevalence of bla _OXA-66 in North Africa and the Middle East (Muthuirulandi Sethuvel et al., 2019) but not with the reported prevalence of bla _OXA-65 in Egypt (Elwakil et al., 2023). A higher prevalence of bla _OXA-51 has been reported in Egypt during the last years, while other variants described in this study have been reported less frequently in Egypt (Elwakil et al., 2023). The genetic contexts of the bla _OXA-51-like variants found in our isolates, were similar to those found in previously described isolates (Vijayakumar et al., 2022). Although ISAba is a commonly found element upstream these genes, not all the variants harbor it necessarily. The most common acquired carbapenemase gene found in these isolates was bla _OXA-23, which is usually found in 90–100% of carbapenem-resistant A. baumannii isolates in Egypt (Abouelfetouh et al., 2019; Hassan et al., 2021). This gene was located within Tn2006 or Tn2008 transposons as previously described, which is crucial for the overexpression and mobilization of bla _OXA-23 (Hamidian and Nigro, 2019). Regarding to bla _GES-like genes, these type of Extended-Spectrum β-Lactamase genes have been increasingly reported during the last years, and some of its variants can possess carbapenemase activity (Bonnin Rémy et al., 2013). These type of β-Lactamase genes are frequently reported in the Mediterranean Area and the Middle East countries such as Turkey, Tunisia, or Kuwait (Bonnin Rémy et al., 2013; Cicek et al., 2014; Chihi et al., 2016). However, in our study, these genes were identified in 25% of the isolates, half of the frequency reported in other studies in Egypt (50%) (Ramadan et al., 2018). As previously described in isolates from Pakistan, bla _GES-like genes were part of a class 1 integron co-harboring dfrA7, sul1 and aac(6’)-Ib resistance genes (Karah et al., 2020). Furthermore, bla _OXA-23 and bla _GES-11 genes have been described encoded in a large conjugative plasmid named pK50a (79.6kb) which is a member of the Aci6 group (Wibberg et al., 2018). This is approximately of the same size as the plasmid in which we identified the bla _GES-11 gene, and it is worth mentioning that all the isolates harboring bla _GES-like genes were positive for replicase Aci6 too. The bla _NDM-1 gene was detected in 27.8% of isolates, in concordance with the literature where NDM-type carbapenemases are commonly reported in Egyptian isolates with a prevalence of 0–39.3% (El-Kholy et al., 2021). This gene was found within the truncated isoform of Tn125 (ΔTn125) in our isolates with the characteristic ISAba125 upstream of the bla _NDM-1 gene which appears to enhance its expression (Wang et al., 2012). In all the isolates, bla _NDM-1 gene was chromosome-borne, which is the most frequent localization (Fernández-Cuenca et al., 2020). The bla _PER-7 gene was located within a complex structure connecting ISCR1 element and class 1 integron, which is a previously described structure closely related to multidrug resistant bacteria (Cheng et al., 2016). However, another ISCR1 element, IS5 and part of IS10A were identified downstream of the 3’-CS which differs from the described structure. NDM-type carbapenemases and PER-type β-lactamases seem to be involved in cefiderocol resistance, the recently developed last resort antibiotic (Naeimi Mazraeh et al., 2021; Poirel et al., 2021).

Analysis of the sequenced genomes showed that seventeen isolates belonged to IC2 with the characteristic bla _OXA-66 gene, which is the dominant clone circulating worldwide and in Egypt (Hassan et al., 2021). Among this lineage, ST570 was the most abundant clade, an ST which has four entries in the PubMLST database, submitted from Vietnam (1) and Egypt (3) indicating that this lineage is circulating in Egyptian health settings at least, since 2017. Regarding to ST2, among 1319 isolates submitted to the database, two entries from Egypt between 2013 and 2015, and eight entries from Jordan between 2019 and 2020 were found. A recent study showed six out of seven Syrian strains pertaining to ST2 (Higgins et al., 2021), demonstrating that ST2 is commonly circulating between countries of the Mediterranean area. The less abundant clade was ST600, which has five entries in the PubMLST database isolated from Jordan (4) and Libya (1), between 2014 and 2020, suggesting a possible transmission between these two nearby countries during these years. Ten isolates belonged to IC5 (ST158) harbouring the characteristic bla _OXA-65 gene. Four isolates were identified in the PubMLST database isolated from Iraq, Turkey, Russia and Egypt; consistent with a study published in 2020 in Egypt, where the majority of the isolates belonged to ST158, indicating that this clone is circulating in the Mediterranean and Middle East area (Elwakil et al., 2023). Three isolates belonged to IC4 (ST15), a ST which is predominantly found in Latin American countries, although it has also been described in countries of the Mediterranean are such as Turkey (Di Popolo et al., 2011). Three isolates were related to IC9 (ST85), which have been recently reported in Libya (Higgins et al., 2021). A search in PubMLST also showed four entries in Jordan during 2020 and one in Egypt in 2017 demonstrating the presence of this clade in Egypt and the Middle East area. Just one isolate belonging to IC7(ST113) was identified, a sequence type frequently reported in South America (Kurihara et al., 2020) although, it has also been described in Cairo in 5 isolates from 2018 to 2020 according to PubMLST. A single isolate pertaining to IC8(ST613) was detected, this is a linage with little presence in the Middle East and North Africa, with just one isolate reported in Alexandria in 2013 according to PubMLST. A singleton assigned to ST164 was identified among our isolates, this ST have been identified in Germany in 2021 (Wareth et al., 2021) and in Turkey in 2016. There are no public records of isolates belonging to this linage in Egypt up to date.

In conclusion, this study showed a high clonal diversity among CRAB isolates collected from hospitals in Alexandria, and highlights the emergence of not frequently reported lineages in Egypt. The high incidence of bla _OXA-23 carbapenemase as well as bla _NDM-1 is of concern as they are key in carbapenem resistance and to many other antibiotics. This work also puts the spotlight in the emergence of cefiderocol resistant isolates in Egyptian hospitals. It becomes necessary to harden infection control measures and to increase epidemiologic studies in Egypt to limit the development of new clones with highly resistant genes.

The datasets presented in this study can be found in online repositories. The names of the repository/repositories and accession number(s) can be found in the article/ Supplementary Material .

SS-U, LG and IA contributed to the design of the experiments and were responsible for the project funding. SS-U, AM-B, AO-S, MH and DA performed the experiments. SS-U, AO-S and LG analyzed and interpreted the data. SS-U and LG wrote the manuscript. SMS and ME-K were responsible for the collection, identification and determination of the resistance profile of the isolates. All authors contributed to the article and approved the submitted version.