REVIEW Minor extended-spectrum b-lactamases T. Naas, L. Poirel and P. Nordmann Service de Bacte´riologie-Virologie, Hoˆpital de Biceˆtre, Assistance Publique ⁄ Hoˆpitaux de Paris, Faculte´ de Me´decine Paris-Sud, Universite´ Paris XI, 94275 K.-Biceˆtre, France

ABSTRACT Extended-spectrum b-lactamases (ESBLs) are usually plasmid-mediated enzymes that confer resistance to a broad range of b-lactams. Initially, resistance to third-generation cephalosporins in Gram-negative rods was mainly due to the dissemination of TEM- and SHV-type ESBLs, which are point mutants of the classic TEM and SHV enzymes with extended substrate specificity. During the last ten years, CTX-Mtype ESBLs have become increasingly predominant, but less frequent class A b-lactamases have also been described, including SFO, BES, BEL, TLA, GES, PER and VEB types. While several of these latter are rarely identified, or are very localised, others are becoming locally prevalent, or are increasingly isolated worldwide. In addition, mutations can extend the spectrum of some OXA-type b-lactamases to include expanded-spectrum cephalosporins, and several of these enzymes are considered to be ESBLs. Extended spectrum b-lactamase, ESBL, non-CTX-M, non-SHV, non-TEM, OXA ESBL, oxacillinase, review

Keywords

Clin Microbiol Infect 2008; 14 (Suppl. 1): 42–52 INTRODUCTION b-Lactamases are most commonly classified according to two general schemes: the Ambler molecular classification and the Bush–Jacoby– Medeiros functional system [1,2]. A commonly used working definition of extended-spectrum b-lactamases (ESBLs) is: b-lactamases capable of conferring bacterial resistance to the penicillins, first-, second-, and third-generation cephalosporins and aztreonam (but not to cephamycins or carbapenems) with hydrolysis rates of at least 10% of that of benzylpenicillin and inhibited by b-lactamase inhibitors such as clavulanate [3]. The extension of the spectrum of some OXA-type b-lactamases towards the expanded-spectrum cephalosporins has also been observed, and several of these enzymes are considered to be ESBLs, even though they are poorly inhibited by clavulanate [3–5]. Although ESBLs have been reported most frequently in Enterobacteriaceae, they have been found in other bacterial species, such as Pseudomo-

Corresponding author and reprint requests: T. Naas, Service de Bacte´riologie-Virologie, Hoˆpital de Biceˆtre, 78 rue du Ge´ne´ral Leclerc, 94275 K.-Biceˆtre, France E-mail: [email protected]

nas aeruginosa and Acinetobacter baumannii [3,4]. For the purpose of this review, the term ESBL will be taken to mean those b-lactamases of Bush–Jacoby– Medeiros group 2be (molecular class A enzymes) and those of group 2d (OXA; molecular class D enzymes), which share most of the properties of group 2be enzymes [3,5]. Initially, the 2be designation indicated that these enzymes are derived from group 2b b-lactamases (such as TEM-1 and SHV-1), with the letter ‘e’ denoting extended-spectrum activity. Subsequently, CTX-M enzymes have also been placed into the 2be group, even though the progenitors of these enzymes are naturally capable of hydrolysing thirdgeneration cephalosporins and aztreonam [3]. While the majority of clinically isolated ESBLs are TEM, SHV or CTX-M types [3,6], several ESBLs that are not closely related to any of the three established families have been reported. Examples include the SFO, BES, BEL, TLA, GES, PER and VEB types (Tables 1 and 2, Fig. 1). These are not simple point-mutant derivatives of any known b-lactamases and, except for SFO-1, their progenitor genes remain unknown. These ESBLs are remarkable for their geographical diversity. This review describes the properties and the distribution of these infrequent class A and class D ESBLs, whether chromosomal- or plasmid-encoded.

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Naas et al. Minor or infrequent ESBLs 43

Table 1. Plasmid-encoded extended-spectrum b-lactamases

b-Lactamase name ‘Old ESBL’ SHV type TEM type ‘New ESBL’ CTX-M type ‘Minor ESBL’ SFO-1 TLA-1 PER VEB BES-1 GES BEL-1 TLA-2 OXA ESBLs OXA

Yeara

No. of variants

Origin of the name

1983 1985

>100 >160

Sulphhydryl variable Patient’s name: Temoneira

1989

>65

1988 1991 1991 1996

1 1 3 5

1996 1998 2005 2005

1 9 1 1

1991

At least 9

Cefotaximase—Munich Serratia fonticola Tlahuicas (Indian tribe) Pseudomonas extended resistance Vietnam extended-spectrum b-lactamase (ESBLs) Brazilian ESBLs Guyana ESBLs Belgium ESBLs 51% amino-acid identity with TLA-1 Hydrolysis of oxacillin > penicillin

a

Year first recorded.

CHROMOSOMAL-ENCODED ESBLs Several Gram-negative species have inherent ESBL genes carried by their chromosome. In these bacteria, the presence of a single b-lactamase gene copy, with a weak promoter, may yield only lowlevel expression of the resistance phenotype [7]. Those Enterobacteriaceae that naturally produce an ESBL are resistant to amino-penicillins, variably resistant to carboxy-penicillins, with this resistance being reversed by inhibitors of class A enzymes such as clavulanate, variably resistant to first- and second-generation cephalosporins, and susceptible to third-generation cephalosporins. Synergy may be observed between first-, secondand sometimes third- or even fourth-generation cephalosporins and clavulanate. This phenotype is specific for a given species, such as Proteus vulgaris (CumA), Proteus penneri (HUGA), Erwinia persicina (ERP-1), Citrobacter sedlakii (SED-1), Kluyvera spp. (KLUA, KLUC, KLUG), Serratia fonticola (FONA), Rahnella aquatilis (RAHN-1) [7–9], or Klebsiella oxytoca (K1 or Koxy) [2,3]. Non-fermentative Gram-negative rods (e.g., Stenotrophomonas maltophilia (L2 enzyme), Chryseobacterium meningosepticum (CME-1 ⁄ 2), Chryseobacterium gleum (CGA-1) and Burkholderia pseudomallei (PENA)) and strictly anaerobic bacteria (Desulfovibrio desulfuricans (DES-1), Prevotella intermedia (CFXA2) and Bacteroides fragilis (CEPA)) have also been shown to possess ESBL genes [10,11].

These chromosomally encoded enzymes may be at the origin of plasmid-encoded enzymes. b-Lactamases of the CTX-M groups are structurally related to the naturally produced b-lactamases of Kluyvera ascorbata (CTX-M-2) Kluyvera georgiana (CTX-M-8), Kluyvera cryocrescens (CTX-M-1), and Kluyvera spp. isolated in Guyana (CTX-M-9) [6]. Similarly, the plasmid-encoded SFO-1 enzyme is highly related to the chromosomally encoded FONA-1 enzyme from S. fonticola [9,12]. A dendrogram of the phylogeny of some chromosomaland plasmid-encoded ESBLs is shown in Fig. 1. PLASMID-ENCODED ESBLs Besides the CTX-M-type enzymes, several other plasmid-mediated ESBLs are not simple pointmutant derivatives of classic b-lactamases. While some of these enzymes have so far been isolated only once or twice, others are increasingly being isolated worldwide (Tables 1 and 2). Rare ESBLs SFO-1. SFO-1 was found in a single clinical Enterobacter cloacae isolate from Japan in 1988 [12]. SFO-1 hydrolyses cefotaxime very efficiently, ceftazidime poorly, and spares cephamycins and carbapenems. Its activity is inhibited by clavulanate and imipenem. An unusual feature of SFO-1, which is highly related to a class A b-lactamase

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44 Clinical Microbiology and Infection, Volume 14, Supplement 1, January 2008

Fig. 1. Phylogeny of some chromosomaland plasmid-encoded extended-spectrum b-lactamases (ESBLs). Plasmid-encoded minor class A ESBLs are boxed and OXA ESBLs are indicated. The dendrogram was constructed using CLUSTALW aligned protein sequences.

from S. fonticola, is that it was encoded on a selftransferable plasmid and that the b-lactamase production was induced to a high level by imipenem [9,12]. The encoding plasmid also carries the ampR regulatory gene necessary for the induction of the b-lactamase in a manner similar to class C b-lactamases. However, unlike class C b-lactamases, SFO-1 cannot hydrolyse cephamycins and is well-inhibited by clavulanate [12]. BES-1. BES-1was isolated once from a Serratia marcescens strain from a hospital in Rio de Janeiro (Brazil) in 1996 [13]. It confers a high level of resistance to aztreonam and a higher level of resistance to cefotaxime than to ceftazidime. Its activity against cefotaxime resembles that of CTX-M types; however, BES-1 is more active against ceftazidime and has a 1000-fold higher affinity for aztreonam.

The blaBES-1 gene is plasmid-encoded and its product belongs to functional group 2be, with 48% amino-acid identity with the CTX-M group 1 b-lactamases. BES-1 is well-inhibited by clavulanate, whereas tazobactam is a poor inhibitor [13]. BEL-1. BEL-1 was identified in a P. aeruginosa strain isolated in Belgium in 2004, which displayed on a disk diffusion antibiogram an image of synergy between clavulanate and ceftazidime containing disks, thus indicating the presence of an ESHL [14]. BEL-1 is weakly related to other Ambler class A ESBLs (50% amino-acid identity with GES-1 ESBL, 40% with CTX-M group 8, 36% with BES-1). BEL1 significantly hydrolyses most expanded-spectrum cephalosporins and aztreonam, and its activity is well-inhibited by clavulanate, cefoxitin, moxalactam, and imipenem, whereas tazobactam is a poor inhibitor, as with BES-1. The blaBEL-1 gene is chromosomally located and is embedded in a

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Naas et al. Minor or infrequent ESBLs 45

Table 2. MICs (mg ⁄ L) for clinical strains, Escherichia coli strains harbouring recombinant plasmids (rec) or E. coli transformant (Tc) strains b-Lactamase

Species

SFO-1a

Enterobacter cloacae Escherichia coli (Tc) Serratia marcescens E. coli (rec) E. coli Pseudomonas aeruginosa Salmonella Typhimurium E. coli (Tc) P. aeruginosa E. coli (Tc) P. aeruginosa E. coli (rec) Klebsiella pneumoniae E. coli (Tc)

BES-1b TLA-1c PER-1d VEB-1e BEL-1f GES-1g

TIC

TCC

PIP

TAZ

>512 >512

512 32

512 512

256 256

>512 >512 >512 >512 256 >512 >512 >512 256

256 128 1 8 4 128 64 64 8

8 256 16 256 16 16 128 512 16

8 64 2 8 2 8 32 64 2

CTX

CAZ

128 8 64 64 >256 256 256 128 >512 2 32 1 0.5 0.5

4 2 4 16 >256 256 512 512 >512 256 32 16 4 8

CAZ-Ac

1 0.25 4

16 1 4 2 0.5 1

FEP

ATM

IPM

4 8 64 32 128 8 256 1 4 0.25 0.25 0.25

16 16 512 512 >256 256 128 512 512 32 32 16 0.12 0.25

1 0.25 0.5 0.25 1 1 0.12 0.06 0.5 0.12 1 0.06 0.12 0.06

a

[12]; b[13]; c[16]; d[19]; e[41]; f[14]; c[52]. TIC, ticarcillin; TCC, ticarcillin ⁄ clavulanate; PIP, Piperacillin; TAZ, Piperacillin/Tazocillin; CTX, Cefotaxim; CAZ, Ceftazidime; CAZ-Ac, Ceftazidime/clavulanatc; FEP, Cefepime; ATM, Aztreonam; IPM, imipenem.

class 1 integron containing three other gene cassettes [14]. Recent work suggests that the blaBEL-1 gene might be disseminating in Belgium in clinical and community P. aeruginosa isolates [15]. TLA-1. TLA-1 was identified in an Escherichia coli isolate from a patient in Mexico in 1993 [16]. It hydrolyses expanded-spectrum cephalosporins, including cefotaxime, ceftazidime, aztreonam and cefepime, but not imipenem and cefoxitin. It is strongly inhibited by tazobactam and, to a lesser extent, by clavulanate and sulbactam. The blaTLA-1 gene is localised on a self-transferable plasmid. Its amino-acid sequence shares 50% identity with CME-1, the chromosomal class A b-lactamase from C. meningosepticum, and c. 40% homology with the plasmid-encoded VEB and PER enzymes. Nosocomial bacteraemia and urinary tract infections caused by Klebsiella pneumoniae with plasmids carrying both SHV-5 and TLA-1 genes have been recently described in Mexico [17]. TLA1 remains unreported outside of Mexico. TLA-2. TLA-2 is encoded by a 47-kb plasmid that was isolated from an unidentified bacterial strain obtained from the activated sludge compartment of a wastewater treatment plant in Germany in 2002 [18]. TLA-2 is an Ambler class A b-lactamase that shares 52% amino-acid identity with CGA-1 from C. gleum and only 51% with TLA-1 from

E. coli. TLA-2 has good catalytic efficiency against most cephalosporins, but not against penicillins, and it lacks sensitivity to b-lactam inhibitors. Although there is no evidence yet for diffusion of TLA-2 into clinical settings, the fact that this enzyme can degrade expanded-spectrum cephalosporins and is encoded by a plasmid that can be mobilised may indicate future clinical relevance. Increasingly isolated ESBLs PER-1. PER-1 was initially detected in 1993 in a P. aeruginosa isolate from a Turkish patient hospitalised in France [19,20]. This enzyme is weakly related to other ESBLs and confers resistance to penicillins, cefotaxime, ceftazidime, and aztreonam but spares carbapenems and cephamycins. Its activity is well-inhibited by clavulanate, sulbactam and tazobactam [19,20]. Although PER-1 is typical among class A enzymes in its catalytic machinery, its three-dimensional structure reveals that it forms a subgroup in the class A superfamily, with a new fold in the W-loop a region, considered to be a canonical motif in class A enzymes [20,21]. Several enzymes with significant sequence identity (40%) were subsequently identified as belonging to the ‘PERlike’ family: PER-2, VEB-1, TLA-1 and chromosomal CME-1 from C. meningosepticum [21]. The blaPER-1 gene is widespread in Acinetobacter spp. and P. aeruginosa, but can also be isolated

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46 Clinical Microbiology and Infection, Volume 14, Supplement 1, January 2008

from Salmonella enterica serovar Typhimurium and Providencia rettgeri in Turkey [22,23]. A recent survey in Turkish intensive care units revealed that, among ceftazidime-resistant nosocomial isolates of Acinetobacter spp. and P. aeruginosa, 32% and 55%, respectively, had the PER-1 enzyme [22]. Although organisms expressing PER-1 have been found predominantly in Turkey, outbreaks of P. aeruginosa with the enzyme have occurred in Italy, with no apparent contacts with Turkish patients [24]. Worryingly, a P. aeruginosa strain producing both PER-1 and the carbapenemase VIM-2 has been detected in Italy [25]. This combination renders the organism resistant to virtually all b-lactam antibiotics. PER-1 has also been found in Proteus mirabilis and Alcaligenes faecalis in Italy [26,27]. A recent survey of ESBLproducing Enterobactericeae conducted in a French university hospital in 2002 and in Spain in 2005 revealed PER-1 in enterobacterial isolates, e.g., Providencia stuartii [28] and P. mirabilis [29]. Isolates of P. aeruginosa and A. baumannii producing PER-1 have been detected on several occasions in France, Belgium and recently in Japan [30–32], and a high prevalence of PER-1 in Acinetobacter spp. from Korea was noted [33]. PER-3, a point-mutant derivative of PER-1, was identified in Aeromonas caviae in France [34]. A related enzyme, PER-2, which has 86% aminoacid homology with PER-1, was found among S. enterica serovar Typhimurium strains in Argentina in 1996 [35] and subsequently in other Gram-negative bacteria, including S. enterica serovar Senftenberg, K. pneumoniae, Enterobacter aerogenes, E. cloacae, Vibrio cholerae and, recently, A. baumannii. It has been reported in Argentina, Uruguay, and Bolivia [36–40]. Whereas PER-1 is reported mostly in Turkey and Korea, PER-2 has only been reported in South America thus far. VEB-1. The blaVEB-1 gene, first reported in 1996 in relation to an E. coli isolate from a Vietnamese patient, is plasmid- and integron-located [41,42]. VEB-1 has highest amino-acid identity with PER-1 and PER2 (38%) [20], and confers high-level resistance to ceftazidime, cefotaxime and aztreonam. VEB-1 is equally well-inhibited by clavulanate, sulbactam and tazobactam, and also by moxalactam, imipenem and cefoxitin, as best evidenced in enterobacterial isolates on disk diffusion antibiogram

by an unusual image of synergy between a cefoxitin- and a cefuroxime-containing disk. The activity against expanded-spectrum cephalosporins is very high in general, except for ceftazidime and aztreonam, whereas the hydrolytic activities against penicillins are much lower [41]. After its initial discovery, the blaVEB-1 gene was detected in two P. aeruginosa isolates from Thailand, where it was chromosomal- and integronlocated [43]. Several epidemiological surveys in Thailand and Vietnam have emphasised the widespread distribution of VEB-1. In these countries, up to 40% and 80% of ceftazidime-resistant enterobacteriacae and P. aeruginosa isolates, respectively, were positive for blaVEB-1 [43–45]. Four single point-mutant derivatives of VEB-1 (VEB-2 to -5) have been described (http://www.lahey. org/Studies/other.asp#table1). In addition, single VEB-producing isolates have been detected in P. aeruginosa isolates from Kuwait, China, India and Bangladesh, in A. baumannii isolates from France, Belgium and Argentina, in P. stuartii isolates from Algeria, in E. cloacae and Achromobacter xylosoxidans isolates from France, and in E. coli isolates from Canada [28,29,40,46,47]. Outbreaks involving isolates with VEB-1 enzymes have been described, including a nationwide outbreak of VEB-1 A. baumannii in France and Belgium [29,48], P. mirabilis in Korea [49] and VEB-3 E. cloacae in China [50]. In most instances, VEB-1 is integron-borne and can be associated with several other resistance genes. In the prototype E. coli producer MG-1, the blaVEB-1 gene was carried on an integron, In53, that contained eight additional gene cassettes, including those coding for the OXA-10 b-lactamase, several aminoglycoside resistance markers, and the rifampin ADP ribosyltransferase, chloramphenicol resistance marker, and one specifying resistance to quaternary ammonium compounds [42]. The blaVEB-1 gene was characterised in a peculiar genetic environment in P. aeruginosa isolates from India and Bangladesh and in P. stuartii from Algeria. Instead of being part of a typical class 1 integron structure, the blaVEB-1 gene is flanked by identical 135-bp sequences, termed repeated elements (Res), which are bracketed by two truncated 3¢-conserved sequences of class 1 integrons in direct repeat [46]. These Re structures carry a strong promoter that drives the expression of the downstream-located

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Naas et al. Minor or infrequent ESBLs 47

blaVEB-1a gene. This is a novel finding, since most ESBL genes have been reported to be associated with either transposons or integrons, but always with a single type of genetic vehicle. The finding of a similar structure on different continents suggests that these Re elements may be widespread and that blaVEB-1 genes may disseminate via different genetic elements [46]. A frequent association of the plasmid-mediated quinolone resistance determinant qnrA and the blaVEB-1 gene has been identified in enterobacterial isolates from France, Turkey, Thailand and Canada [51]. This association may explain, in part, the frequent co-resistance to quinolones and expanded-spectrum b-lactams in isolates producing VEB-1 b-lactamase. GES. ESBLs of the GES type (also named IBC) are reported increasingly in Gram-negative rods, including P. aeruginosa, E. coli and K. pneumoniae [52,53]. The current GES nomenclature is available via the Lahey Clinic website (http:// www.lahey.org/studies/webt.html). GES-1 was initially characterised in a K. pneumoniae isolate from France, where it was plasmid- and integronencoded [52]. GES-2 was from South Africa, GES5, GES-6, GES-7 and GES-8 were from Greece, and GES-3 and GES-4 were from Japan (Table 3; [54]. GES-5 has also been reported recently from Korea, China and Brazil [55,56]. Thus, similar GES variants have now been identified in widely separated countries (Table 3 [54]). GES-1 possesses a hydrolysis profile similar to that of classic clavulanate-inhibited Ambler class A ESBLs, including activity against penicillins and expanded-spectrum cephalosporins, but not cephamycins and carbapenems, and is inhibited by clavulanate, tazobactam and imipenem [52]. Unlike most ESBLs, GES-1 does not hydrolyse aztreonam. GES-2 also hydrolyses carbapenems and is less susceptible to inhibitors, due to a 2-bp substitution (Table 3) (Fig. 1), leading to a single Gly170Asn change inside the W-loop of the catalytic site [57]. A Gly170Ser change was identified in GES-4, GES-5, and GES-6 (Table 3), resulting in slow carbapenem hydrolysis, weak inhibitor susceptibility (as for GES-2), but, additionally, cephamycin hydrolysis, as shown for GES-4 [58]. GES-9, which differs from GES-1 by a Gly243Ser change (Table 3), does not hydrolyse

carbapenems but has broadened activity against monobactams [59]. Worryingly, P. aeruginosa strains producing GES-1 and the carbapenemase VIM-11 and E. coli strains producing GES-7 and the carbapenemase VIM-2, encoded on separate integrons, have been detected in Greece and in Argentina, respectively, and are virtually pan-resistant isolates [60,61]. Several outbreaks of Gram-negative bacteria producing GES variants have been described: K. pneumoniae in Korea, Portugal and Greece; S. marcescens in The Netherlands; and P. aeruginosa in South Africa [59, 62–66]. GES-2 is the first described example of an ESBL that extended its spectrum of activity against carbapenems by a single point mutation. Subsequently, four other variants with similar carbapenemase activity have been described (GES-4, GES5, GES-6 and GES-8). This may represent an intermediate step between ESBLs and class A carbapenem-hydrolysing enzymes such as KPC. In fact, KPC-type enzymes possess the properties required for being called ESBLs, but are also capable of significant carbapenem hydrolysis [67]. OXA ESBLs Although the definition of ESBLs is often restricted to class A enzymes, several oxacillinases with extended-spectrum activity may be also included, and are called OXA ESBLs [3–5]. In general, the OXA b-lactamases (group 2d) are so named because of their oxacillin-hydrolysing abilities, with hydrolysis rates for cloxacillin and oxacillin being greater than 50% that for benzylpenicillin [5]. They predominantly occur in P. aeruginosa but have been detected in many other Gram-negative bacteria, including Enterobacteriaceae [5]. Most OXA-type b-lactamases do not significantly hydrolyse the expanded-spectrum cephalosporins, and the evolution of OXAtype ESBLs from parent enzymes with narrower spectra has many parallels with the evolution of SHV- and TEM-type ESBLs. While most OXAtype ESBLs derive from OXA-10 (OXA-11, OXA14, OXA-16 and OXA-17) or OXA-13, which is a ten-amino-acid derivative of OXA-10, (OXA-19 and OXA-28) or to a lesser extent from OXA-2 (OXA-15 and OXA-32), others are unrelated to any recognised broad-spectrum OXA enzymes, e.g., OXA-18 and OXA-45 [3,5,68–70].

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AF156486

AF326355 AB113580 AB116260 AY494717

AY494718 AF208529 AF329699 AY920928

GES variant

GES-1

GES-2 GES-3 GES-4 GES-5

GES-6 GES-7 (IBC-1) GES-8 (IBC-2) GES-9

France, Argentina, Brazil, Portugal and The Netherlands South Africa Japan, China and Korea Japan Greece, Korea, China and Brazil Greece Greece Greece France

Country

b

Thr Thr

Lys Lys

Lys Lys

Glu

Leu

Ala

Ser

Ser Ser

Asn

Gly

Ser

Gly

+ + + +

+ + + +

+

+ ) + )

) ) ) + + ) ) )

+ ) + +

) ) ) )

) ) + +

)

)

)

IPM

ATM

FOX

Met

243

Klebsiella pneumoniae, Pseudomonas aeruginosa, Serratia marcescens P. aeruginosa K. pneumoniae K. pneumoniae Escherichia coli, K. pneumoniae, P. aeruginosa K. pneumoniae Enterobacter cloacae P. aeruginosa P. aeruginosa

170

CAZ

126

62

Strain

104

Hydrolysis profileb

Ambler positiona

Only amino-acid changes as compared to GES-1 are indicated. + and –, hydrolysis and no hydrolysis, respectively. c p means poorly inhibited and s means similarly inhibited as compared to GES-1. CAZ, Ceftazidime; FOX, cefoxitin; ATM, Aztreonam; IPM, imipenem; Ac clav.

a

GenBank accession numbers

Table 3. Key amino-acid substitutions of GES variants in relation to their hydrolytic spectra

P S S S

P S P P

S

Ac clav

Inhibitionc

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Naas et al. Minor or infrequent ESBLs 49

OXA-10 and OXA-13 themselves weakly hydrolyse cefotaxime, ceftriaxone and aztreonam, thus providing most organisms with reduced susceptibility to these antibiotics. Most OXA10-derived ESBLs confer greater resistance to cefotaxime, and significant resistance to ceftazidime and aztreonam, except for OXA-17, which has little effect on the MICs of ceftazidime, but has substantial activity against cefotaxime [4,5]. OXA ESBLs exhibit appreciable diversity in their enzymic activities. Although most OXA-type enzymes are resistant to b-lactamase inhibitors, OXA-18 and OXA-45 have been reported to be very well inhibited by clavulanate [69,70]. There are very few epidemiological data on the geographical spread of OXA-type ESBLs. The OXA-type ESBLs were originally discovered in P. aeruginosa isolates from a single hospital in Ankara, Turkey [68]. These were mostly OXA-10 derivatives, and a recent survey revealed that they were present in 55% of ceftazidime-resistant P. aeruginosa isolates from intensive care units in Istanbul, Turkey [23]. OXA ESBLs have also been characterised in 2.9% and in 0.4% of ceftazidimeresistant P. aeruginosa isolates in Taiwan and in Korea, respectively [71,72]. In France, single novel derivatives of OXA-13 (OXA-19 and OXA-28) and OXA-18 were found in a few P. aeruginosa isolates, and OXA-45 was found in one P. aeruginosa isolate from Texas, USA [5,69,70]. CONCLUSION Numerous chromosomally encoded ESBLs have been described worldwide, and it is likely that many more will be discovered in the future. These enzymes may represent potential threats, since gene-capture units may mobilise them onto plasmids and may enhance their dissemination. Several of these genes have already been mobilised, with CTX-M types being very successful. Others, SFO-1, BES-1, BEL-1, TLA-1 and TLA-2, are as yet very rare and are geographically localised, whereas yet others, GES, PER and VEB, have been described on several continents. In many cases, extensive epidemiological surveys have not yet been undertaken, and the precise distribution is not known. Identification and knowledge of distribution may more likely reflect the interest of research teams than true prevalence. With ESBLs, the saying ‘we find only what we are looking for’ is very appropriate.

Another worrying aspect is that some of these enzymes may extend their spectrum of hydrolysis towards carbapenems by simple point mutations, as illustrated by GES-2. Furthermore, the number of point-mutant derivatives of GES variants and their geographical spread signals a worrying ongoing evolution in this family of enzymes. Unlike the genes of the TEM-, SHV- and CTXM-type enzymes, those of most minor ESBLs are carried by integrons in association with other resistance genes (mainly aminoglycoside resistance genes), thus ensuring co-resistance, coexpression and co-selection of a multiresistance phenotype. Furthermore, as shown for the blaVEB-1 gene, a frequent association with plasmid-encoded rifampicin and quinolone resistance genes may be observed [43,51]. Minor ESBLs, although currently very rare and geographically localised, may rapidly become major concerns, especially since regional occurrence may rapidly be amplified through transcontinental transfer of resistance genes, as now exemplified repeatedly [29,48]. This scenario highlights the difficult problem of screening patients for carriage of multidrug-resistant isolates upon hospital entry. REFERENCES 1. Ambler RP, Coulson AF, Frere J-M et al. A standard numbering scheme for the class A b-lactamases. Biochem J 1991; 276: 269–270. 2. Bush K, Jacoby GA, Medeiros AA. A functional classification scheme for b-lactamases and its correlation with molecular structure. Antimicrob Agents Chemother 1995; 39: 1211–1233. 3. Bradford PA. Extended-spectrum b-lactamases in the 21st century: characterization, epidemiology, and detection of this important resistance threat. Clin Microbiol Rev 2001; 14: 933–935. 4. Paterson DL, Bonomo RA. Extended-spectrum b-lactamases: a clinical update. Clin Microbiol Rev 2005; 18: 657– 686. 5. Naas T, Nordmann P. OXA-type b-lactamases. Curr Pharm Des 1999; 5: 865–879. 6. Pitout JD, Nordmann P, Laupland KB, Poirel L. Emergence of Enterobacteriaceae producing extended-spectrum b-lactamases (ESBLs) in the community. J Antimicrob Chemother 2005; 56: 52–59. 7. Vimont S, Poirel L, Naas T, Nordmann P. Identification of a chromosome-borne expanded-spectrum class a b-lactamase from Erwinia persicina. Antimicrob Agents Chemother 2002; 46: 3401–3405. 8. Petrella S, Clermont D, Casin I, Jarlier V, Sougakoff W. Novel class A b-lactamase Sed-1 from Citrobacter sedlakii: genetic diversity of b-lactamases within the Citrobacter genus. Antimicrob Agents Chemother 2001; 45: 2287–2298.

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