The World Health Organization (WHO) and CDC reported the current and future threat of infections by antimicrobial-resistant microorganisms with a high level of concern [1,31]. Carbapenem-resistant Acinetobacter baumannii (CRA), carbapenem-resistant Pseudomonas aeruginosa (CRP), and carbapenem-resistant Enterobacterales (CRE) have been listed as critical priority pathogens by the WHO [31]. In addition, CRE and CRA have been reported as the most urgent threats by the CDC since 2019 [1]. Particularly, CRE results in 1100 deaths and 13,100 infections in the USA [1], with a high fraction of these infections potentially resulting in death due to limited antibiotic therapies [1,22,30].

Carbapenems, a broad-spectrum β-lactam antibiotic, are structurally related to penicillin [32]. Carbapenems have a carbon instead of a sulfone at the fourth position of the β-lactam ring, differing from other β-lactams. The unique structure plays a major role in their stability against β-lactamases [33]. Carbapenems are not easily diffusible through the cell wall, but they enter the bacteria through outer membrane proteins (porins). Then, carbapenems degrade the cell wall at the penicillin-binding proteins (PBPs) via the β-lactam ring. The mode of action weakens the glycan backbone in the cell wall due to autolysis, and the cell is destroyed because of osmotic pressure [32,33,34].

Carbapenems have been used as last-line agents against Gram-negative, Gram-positive, and anaerobic bacteria [33]. The last-resort antibiotics were approved for clinical use in humans and released into the market in 1985 [1,35,36]. Carbapenems may occasionally be used for pets under certain conditions, according to the Animal Medicinal Drug Use Clarification Act (AMDUCA) [37,38]. Among carbapenems, ertapenem and panipenem have limited use against non-fermentative Gram-negative bacteria but are appropriate for community-acquired infections. Carbapenems, including imipenem, meropenem, doripenem, and biapenem, have been widely used in hospital-acquired infections. These carbapenems are typically reserved for use in patients infected with multi-drug resistant (MDR) bacteria, including extended-spectrum β-lactamase (ESBL)-producing and ampicillinase C (AmpC)-producing bacterial infections [22,23,34], such as complicated intraabdominal and urinary infections, bloodstream and skin infections, community-acquired and nosocomial pneumonia, meningitis, and febrile pneumonia [35,39,40].

Carbapenem-resistant bacteria were first described in 1996 with the identification of carbapenemase-producing Klebsiella pneumoniae [11,23]. In the last decade, the emergence and spread of carbapenem-resistant bacteria have globally increased. For example, many infections caused by CRE are mostly seen in patients in hospitals, long-term care facilities, and long-term acute care hospitals [41,42,43]. Such infections are high risk for patients using ventilators, urinary catheters, intravenous catheters, and long-term antibiotic treatment and for immunocompromised patients [41]. A significant fraction of these infections result in death due to limited treatment options [1,22,30,44]. Specifically, bloodstream infection by CRE causes a high mortality rate in pediatric populations [40,45]. The characteristics, mechanisms, and outcomes of carbapenem-resistant bacteria are thus crucial to prevent and manage such infections [46].

CRE are Enterobacterales resistant to at least one of the carbapenem antibiotics based on their antibiotic susceptibility profile (phenotypic definition) [41]. There are different mechanisms (e.g., genotypic); carbapenem resistance mainly develops when bacteria (1) acquire structural changes in penicillin-binding proteins (PBPs), (2) show a decrease or loss of specific outer membrane porins that filter carbapenems from reaching their site of action, (3) activate the efflux pumps to remove the antibiotics and regulate the intramembrane environment, and (4) acquire β-lactamases and carbapenemases to degrade or hydrolyze carbapenems and other β-lactam antibiotics (e.g., penicillins and cephalosporins) [32,33,34,41]. In addition, carbapenem resistance can be acquired by a combination of CTX-M (activity against cefotaxime) and AmpC enzymes, allowing low-level carbapenem resistance. Further, the combination of the β-lactamase expression and porin gene mutations is associated with high-level carbapenem resistance, attenuating therapy responses [47].

Overall, CRE can become resistant through chromosomal mutations in the porin gene (non-carbapenemase-producing CRE) and/or the production of carbapenem hydrolyzing-enzymes (carbapenemase-producing (CP) CRE) [41]. The presence or expression of the gene coding carbapenemase is usually sufficient for carbapenem resistance, covering 30% of CRE. Thus, CP-CRE is a subset of all CRE [22,41]. These genes are often on mobile genetic elements, leading to their rapid spread and resulting in infections and colonization [1,14,41,48]. Many CRE-colonized individuals do not develop infections; however, they can still spread the bacteria [41]. Similarly, the transfer of genetic elements can occur in the food chain and the environment [1,14,41]. Therefore, routine tests for these carbapenemases through the Antibiotic Resistance Laboratory Network and CDC laboratories are conducted to prevent and control their emergence and spread [41].

A large variety of carbapenemases have been classified into three groups: Ambler Classes A, B, and D β-lactamases, based on hydrolytic and inhibitor profiles using active catalytic substrates of serine or zinc [13,23,32,34,49]. The characteristics of the three most common classes of carbapenemases are detailed and listed in Table 1 [23,49].

Class A enzymes, serine β-lactamases, hydrolyze a broad variety of β-lactam antibiotics, including carbapenems, cephalosporins, penicillin, and aztreonam [49]. These enzymes were identified as chromosomally encoded and plasmid-encoded types [49]. Some of the chromosomally encoded genes are NmcA (not metalloenzyme carbapenemase A), SME (Serratia marcescencens enzyme), IMI-1 (imipenem hydrolyzing β-lactamase), and SFC-1 (Serratia fonticola carbapenemase-1). The plasmid-encoded genes are KPC (Klebsiella pneumoniae carbapenemase), IMI (Imipenem-hydrolyzing beta-lactamase), and GES (Guiana extended spectrum) [23,32]. Among these, the KPC type is the most prevalent enzyme and causes outbreaks in many Asian, African, North American, and European countries [23,32]. KPC gene is mainly located within a 10-kb length, mobile transposon Tn4401, frequently established on conjugative plasmids. The link of bla_KPC with plasmids and transposons assists in intraspecies gene transfer and the dissemination of the gene [50]. Several KPC variants have rapidly increased, and 84 KPC alleles have been recorded in the GenBank database [51]. Of these, KPC-2 and -3 are the most common enzymes worldwide, and 22 KPC variants have also conferred ESBL-, CTX-M-, or ceftazidime-avibactam (CZA)-resistance in their gene position. For example, the KPC-2 gene was carried on the NTE_KPC-Ib transposon on plasmids with a 15-bp insertion, which also harbored the resistance gene, CZA resistance [47,51]. Overall, KPC types are mostly found in Klebsiella pneumoniae, Klebsiella oxytoca, E. coli, and Serratia marcescens, as well as in Enterobacter, Salmonella, and Proteus species [13,23,32]. Their rapid spread and diverse variants severely threaten human health and impact therapeutic efficacy [13,32,51].

Class B enzymes are known as Metallo-β-lactamases (MBL) since they utilize metal ions (usually Zinc) as a cofactor to attack the enzyme’s active site (β-lactam ring). There are 10 types of MBLs; the most important ones include New Delhi Metallo-beta-lactamase (NDM), Verona Integron-Encoded Metallo-beta-lactamase (VIM), and Imipenemase (IMP) [23,32,41,52]. They hydrolyze all current β-lactam antibiotics, except for monobactams (e.g., aztreonam) [53]. IMP was first reported in Japan in S. marcescens in the early 1990s [13], and over 85 sequence variants have been described [53]. IMP variants are found in Acinetobacter and Pseudomonas species, as well as in the Enterobacteriaceae family [13,32]. VIM was then identified in P. auregionasa in Verona, Italy, in late 1997, and over 69 variants have been described [53]. VIM variants are mostly found in Pseudomonas, Acinetobacter, and Enterobacteriaceae species, which are globally distributed [13,32,53]. Recently, NDM was the most prevalent MBL, first identified in Klebsiellea pneumoniae and E. coli isolated from a patient who traveled from India to Sweden in 2008 [13,53]. There have been 29 NDM variants described, and NDM-1 is the most prevalent type. NDM variants are generally dominant in Klebsiella pneumoniae, E. coli, Acinetobacter baumannii, and Pseudomonas aeruginosa [32,52,53].

Class B enzymes are usually found in plasmid vectors or other mobile genetic elements [49]. For instance, IMP and VIM are mostly integron-associated; they are encoded by gene cassettes within class 1 or 3 integrons that may be embedded in transposons, allowing insertion into the bacterial plasmids [53]. NDM is not integron-associated; it has been observed in plasmids rapidly disseminated worldwide [52,53]. Additionally, NDM-producing bacteria can have both NDM-1 and a type IV secretion system (T4SS) gene cluster in plasmids, showing high virulence [52]. Further, NDM-producing bacteria may harbor other carbapenemases in plasmids (e.g., KPC, VIM, and OXA types) and ESBLs [13,41,47,52]. Thus, the emergence of NDM-producing bacteria with increasing variants is a significant threat to public health.

Class D enzymes, serine β-lactamases, are oxacillinase or oxacillin-hydrolyzing enzymes (OXA), comprising over 200 enzymes. OXA rapidly mutates and expands its spectrum activity; the most prevalent carbapenem-hydrolyzing enzymes are OXA-48 and OXA-181 in over 40 carbapenemase variants [36]. OXA-48 was first identified in Klebsiella pneumoniae in Turkey in 2001 [54,55]. Plasmids are the primary genetic elements for the transmission and propagation of the genes; the most frequent hosts for OXA-48 are self-conjugative 60- to 70-kb plasmids [55]. Currently, OXA-48 and OXA-101 variants are mostly dominant in Klebsiella pneumoniae in Turkey, the Middle East, North Africa, and Europe [13,32,36,55]. However, it should be noted that OXA-producing bacteria often have low-level resistance due to weak expression, which is risky for false positive detection and suitable treatment options [55].

The genes coding carbapenemase in β-lactamase (bla) are defined as bla_KPC, bla_NDM, bla_OXA-48, bla_VIM, and bla_IMP [13,34,56]. These genes are found in many bacteria, such as E. coli, K. pneumoniae, Salmonella, Acinetobacter, and Pseudomonas. These bacteria are isolated not only from humans but also animals, food supplies, and water sources worldwide [23,48,57], detailed in the next section.

Several studies have shown that healthcare settings can lead to the spread of CP pathogens in humans [23,29,58]. Frequent hospital visits and long-term stays in healthcare facilities represent a high risk of colonization and infection development with CP bacteria, particularly with CP-CRE [23,29]. For instance, KPC-producing K. pneumoniae caused hospital outbreaks in many European countries such as Greece, Italy, Spain, France, and Germany [59,60,61]; NDM and KPC-producing K. pneumoniae were identified in transplanted patients in Brazil [62]; CP-CRE were found to spread in hospital and community settings in Africa [63,64,65] and Asia [58,66,67]. Another factor of CP-CRE spread is international travel and medical tourism [23]. For instance, KPC-producing K. pneumoniae and E. cloacae were isolated from patients in New York who had recently traveled from France and Greece [68,69]. In another example, NDM-producing K. pneumoniae and E. coli were isolated from Sweden and UK patients who recently traveled to India [70,71].

Among CP-CRE, E. coli and K. pneumoniae have been disseminated globally at an alarming rate in the medical community as critical human pathogens [72,73]. For instance, KPC-producing K. pneumoniae has been found in more than 100 different sequence types (STs). Particularly, K. pneumoniae ST258 is predominant and primarily associated with KPC-2 and KPC-3 production. ST258 comprises two distinct lineages, clades I and II, and ST258 is a hybrid clonal complex created by a large recombination event between ST11 and ST442 [73]. Further, ST11, ST340, and ST512 are single-locus variants of ST258 and harbor carbapenemases. ST11 is closely related to ST258, which is associated with KPC, NDM, VIM, IMP, and OXA-48 production [73].

Further, carbapenem resistance in pathogenic E. coli is a major concern because of limited therapy. For instance, E. coli ST131, causing severe urinary infections, has been linked to the rapid global increase in AMR among E. coli strains [72]. Further, FimH30 lineage and virotype C are the common lineage among ST131, contributing to the spread of ST131 associated with carbapenemases. ST131 is most likely responsible for the global distribution of E.coli with KPC, NDM, and OXA-48 production [72]. These sequence types of E. coli and K. pneumoniae pose a major threat to public health because of their worldwide distribution [72,73].

Additionally, hospitals or health-care settings are a reservoir for CP bacteria. Carbapenem residues in human excreta can get into hospital sewage. Due to the selection of a low concentration of antibiotics, bacteria in hospital effluent can become resistant to carbapenems [23]. Hospital sewage may act as a reservoir for resistance genes, where bacteria likely acquire resistance through HGT [23,29]. Likewise, antibiotic residues and resistant genes released into municipal wastewater could contribute to the selection of CRE and their dissemination to ground and surface water, spreading them to the environment [23]. For example, CP E. coli, E. cloacae, K. pneumoniae, and Citrobacter freundii were found in the river and hospital sewage in Portugal [74], China [75], Vietnam [76], and Australia [77,78]. VIM- and KPC-producing E. coli were found in seven waste water treatment plants in the USA [79]; OXA-48 carrying CRE in tap water was found in six states in the USA [78]. In addition, KPC-producing Salmonella was found in human feces, hospital sewage, and effluent in the USA and Brazil [80,81].

Another possible way of CP bacteria transmission to animals and farms is through direct contact with colonized hosts (human and animal) and a contaminated environment (surface water, ground water, soil) [23]. CP bacteria (E. coli, K. pneumoniae, Salmonella, Acinetobacter, Pseudomonas) have been detected in farm animals, poultry, fish, mollusks, and wild birds and animals [23,48,57,82,83,84,85,86,87,88]. The transmission of CP bacteria also alerts food safety, particularly CRE in the food-chain. For instance, CP bacteria were isolated in meat (beef, chicken, pork), seafood (clam, fish, prawn), and vegetables (lettuce, spinach, Chinese cabbage, roselle) [42,89,90,91,92,93,94]. These studies showed major carbapenemases (NDM, VIM, and KPC) present in foods. The presence of CP bacteria in the food chain mainly contributes to their spread worldwide due to the global food trade, posing a risk to human health [93].

Various environmental, microbiological, and clinical investigations have shown that CP-CRE can widely spread in the community, animal and agricultural products, and the environment [23,38,83,95,96,97]. For the early detection and optimal management of the spread and emergence of CRE, some recommendations include (1) the necessity of screening and rapid diagnostic tools for patients who may have visited countries or hospitals with frequent infection by CRE, (2) specific policies and prioritizing funding for the control and management of infections by CRE, (3) clear strategies indicating the use of carbapenems, and (4) international co-operation to reduce the global spread of CRE [98]. As infections caused by particularly CRE are a global concern, the rapid detection of the causative bacteria is of utmost importance [38,46,98].