In e na ional Jou nal o An imic obial Agen s 63 (2024) 107150
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In e na ional Jou nal o An imic obial Agen s
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Sho Communica ion
Ac i i y o cefide ocol and inno a i e β-lac am/ β-lac amase inhibi o
combina ions agains isogenic s ains o Esche ichia coli exp essing
single and double β-lac amases unde high and low pe meabili y
condi ions
Tania Blanco-Ma ín
1 , #
, Isaac Alonso-Ga cía
1 , #
,Lucía González-Pin o
1 , #
,
Michelle Ou eda-Ga cía
1
,Paula Guija o-Sánchez
1
, Inmaculada López-He nández
2 , 5
,
Ma ía Pé ez-Vázquez
3 , 5
, Belén A acil
3 , 5
, Lo ena López-Ce e o
2 , 5
,Pablo F aile-Ribo
4 , 5
,
An onio Oli e
4 , 5
,JuanCa los Vázquez-Ucha
1 , 5
, Alejand o Becei o
1 , 5 , ∗, Ge mán Bou
1 , 5 , †
,
Jo ge A ca-Suá ez
1 , 5 , †
, On behal o he GEMARA/SEIMC-CIBERINFEC S udy G oup on he
ac i i y and esis ance mechanisms o new β-lac ams and β-lac amase inhibi o s
(PROTECT)
1
Se icio de Mic obiología and Ins i u o de In es igación Biomédica A Co uña (INIBIC), Complexo Hospi ala io Uni e si a io A Co uña, A Co uña, Spain
2
Labo a o io de Re e encia pa a ipado molecula y de ección de mecanismos de esis encia a an imic obianos de Andalucía (PIRASOA). Unidad de Ges ión
Clínica de Mic obiología y En e medades In ecciosas, Hospi al Uni e si a io Vi gen Maca ena, Se illa. Ins i u o de Biomedicina de Se illa (IBIS), CSIC,
Uni e sidad de Se illa, Spain
3
Labo a o io de Re e encia e In es igación en Resis encia a An ibió icos e In ecciones elacionadas con la Asis encia Sani a ia, Cen o Nacional de
Mic obiología, Ins i u o de Salud Ca los III, Majadahonda, Mad id, Spain
4
Se icio de Mic obiología and Unidad de In es igación, Hospi al Uni e si a io Son Espases e Ins i u o de In es igación Sani a ia Illes Balea s (IDISBA),
Palma de Mallo ca, Spain
5
CIBER de En e medades In ecciosas (CIBERINFEC), Ins i u o de Salud Ca los III, Mad id, Spain
a i c l e i n o
A icle his o y:
Recei ed 3 Janua y 2024
Accep ed 15 Ma ch 2024
Edi o : S e ania S e ani
Keywo ds:
β-lac amases
Cefide ocol
β-lac am/ β-lac amase inhibi o
combina ions
Pe meabili y
Double-ca bapenemase
Esche ichia coli
a b s a c
Objec i es: To analyse he impac o he mos clinically ele an β-lac amases and hei in e play wi h
low ou e memb ane pe meabili y on he ac i i y o cefide ocol, ce azidime/a ibac am, az eonam/
a ibac am, ce epime/enme azobac am, ce epime/ anibo bac am, ce epime/zidebac am, imipenem/
elebac am, me openem/ abo bac am, me openem/xe ubo bac am and me openem/nacubac am agains
ecombinan Esche ichia coli s ains.
Me hods: We cons uc ed 82 E. coli labo a o y ans o man s exp essing he main β-lac amases ci cu-
la ing in En e obac e ales (70 exp essing single β-lac amase and 12 p oducing double ca bapenemase)
unde high ( E. coli TG1) and low ( E. coli HB4) pe meabili y condi ions. An imic obial suscep ibili y es ing
was de e mined by e e ence b o h mic odilu ion.
Resul s: Az eonam/a ibac am, ce epime/zidebac am, cefide ocol, me openem/xe ubo bac am and
me openem/nacubac am we e ac i e agains all E. coli TG1 ans o man s. Imipenem/ elebac am,
me openem/ abo bac am, ce epime/ anibo bac am and ce epime/enme azobac am we e also highly ac-
i e, bu uns able agains mos o MBL-p oducing ans o man s. Combina ion o β-lac amases wi h
po in deficiency ( E. coli HB4) did no significan ly a ec he ac i i y o az eonam/a ibac am, ce-
epime/zidebac am, cefide ocol o me openem/nacubac am, bu limi ed he e ec i eness o he es o
ca bapenem- and ce epime-based combina ions. Double-ca bapenemase p oduc ion esul ed in he loss
o ac i i y o mos o he compounds es ed, an e ec pa icula ly e iden o hose E. coli HB4 ans o -
man s in which MBLs we e p esen .
∗Co espondence o: Alejand o Becei o, Mic obiology depa men o he Uni e si y Hospi al A Co uña (CHUAC), Ins i u e o Biomedical Resea ch o A Co uña (INIBIC). As
Xubias, 15006, A Co uña, Spain. Tel.: + 34 981178000.
E-mail add ess: alejand o.becei o.casas@se gas.es (A. Becei o) .
# Tania Blanco-Ma ín, Isaac Alonso-Ga cía and Lucía González-Pin o con ibu ed equally o he s udy.
† Jo ge A ca-Suá ez and Ge mán Bou con ibu ed equally o he s udy as senio au ho s.
h ps://doi.o g/10.1016/j.ijan imicag.2024.107150
0924-8579/© 2024 The Au ho s. Published by Else ie L d. This is an open access a icle unde he CC BY-NC-ND license ( h p://c ea i ecommons.o g/licenses/by-nc-nd/4.0/ )
T. Blanco-Ma ín, I. Alonso-Ga cía, L. González-Pin o e al. In e na ional Jou nal o An imic obial Agen s 63 (2024) 107150
Conclusions: Ou findings highligh he p omising ac i i y ha cefide ocol and new β-lac am/ β-lac amase
inhibi o s ha e agains ecombinan E. coli s ains exp essing widesp ead β-lac amases, including when
hese a e combined wi h low pe meabili y o o he enzymes. Az eonam/a ibac am, cefide ocol, ce-
epime/zidebac am and me openem/nacubac am will help o mi iga e o some ex en he u gency o
new compounds able o esis MBL ac ion, al hough NDM enzymes ep esen a g owing challenge agains
which d ug de elopmen e o s a e s ill needed.
© 2024 The Au ho s. Published by Else ie L d.
This is an open access a icle unde he CC BY-NC-ND license
( h p://c ea i ecommons.o g/licenses/by-nc-nd/4.0/ )
1. In oduc ion
The ea men o in ec ions caused by ESBL and
ca bapenemase-p oducing En e obac e ales is becoming an ala m-
ing conce n [1] . The app o al o ce azidime/a ibac am in 2015
pa ly alle ia ed he sho age o agen s a ailable o comba hese
s ains, bu ce azidime/a ibac am esis ance has been inc easingly
desc ibed. The mos common esis ance mechanism in ol es mu-
a ion o he KPC enzyme (e.g., L169P, D179Y), leading o s uc u al
ea angemen ha a ec he ca aly ic cen e o KPC a chi ec-
u e and accele a e ce azidime u no e a es [2] . In addi ion,
ce azidime/a ibac am does no p o ide he apeu ic co e agains
inc easingly encoun e ed me allo- β-lac amase (MBL)-p oducing
s ains. In his con ex , new an imic obial agen s a e u gen ly
needed.
Cefide ocol, imipenem/ elebac am, and me openem/
abo bac am ep esen ecen ly app o ed op ions wi h high
ac i i y agains En e obac e ales. Cefide ocol is a newly de el-
oped cephalospo in wi h enhanced bac e ial pene a ion ia
side opho e-media ed pa hways and inc eased s abili y agains
mos β-lac amases ound in G am-nega i e bac e ia [3] . On he
o he hand, imipenem/ elebac am and me openem/ abo bac am
a e espec i ely new ca bapenem/diazabicyclooc ane (DBO) and
ca bapenem/bo onic acid (BOR)-de i ed β-lac am/ β-lac amase
inhibi o combina ions, ha exhibi p omising ac i i y agains
En e obac e ales p oducing class A o C β-lac amases, pa ic-
ula ly KPC enzymes [ 4 , 5 ]. Un o una ely, esis ance o hese
agen s has al eady been epo ed [6–8] We a e now wi ness-
ing he eme gence o some p omising β-lac am/ β-lac amase
inhibi o s. Az eonam/a ibac am and h ee ce epime-based com-
bina ions (ce epime/enme azobac am, ce epime/ anibo bac am
and ce epime/zidebac am) a e a he la e s age o de elopmen .
Az eonam/a ibac am akes ad an age o he pa icula p ope y
o az eonam o e ade hyd olysis by MBL in combina ion wi h
he high po ency o a ibac am agains se ine ype enzymes, hus
i ually co e ing all ypes o β-lac amases [9] . Enme azobac am
includes a new azobac am de i a i e wi h enhanced ac i i y
agains ESBLs. Tanibo bac am is an inno a i e BOR- ype inhibi o
which p o ec s ce epime om hyd olysis ia class A, C, D, and
some class B enzymes; and zidebac am (DBO) po en ia es he
ac i i y o ce epime h ough syne gis ically imp o ed bac e ial
killing due o i s in insic an i-PBP-2 ac i i y [10] . Finally, he e
a e ea ly-s age inhibi o s in he pipeline: nacubac am, a new DBO-
ype β-lac am enhance wi h an i-PBP-2 ac i i y ha po en ia es
he efficacy o me openem agains ca bapenem- esis an s ains;
[11] and xe ubo bac am, a no el BOR de i a i e ha has been
shown o es o e he ac i i y o me openem agains all ypes o
enzymes (including MBLs) [12] .
These ea men s may ep esen a s ep o wa d in he figh
agains G am-nega i e in ec ions. Howe e , he specific ac i i y o
hese agen s agains he main enzymes ci cula ing in En e obac-
e ales has no ye been ho oughly in es iga ed. Mo eo e , how
addi ional ac o s ha ex end beyond he p oduc ion o single
β-lac amases may limi he e ec i eness o hese new agen s,
such as po in deficiency o simul aneous p oduc ion o mul i-
ple enzymes (a phenomena commonly encoun e ed in high- isk
clones o c i ical p io i y ESKAPE a ge s), has no been analysed
in de ail. He e, we cons uc ed an isogenic panel o E. coli isola es
exp essing single β-lac amases o double ca bapenemases unde
low and high pe meabili y condi ions o e alua e he ac i i y o
hese inno a i e agen s, accu a ely de e mine hei he apeu ic
niches, and an icipa e hei po en ial esis ance mechanisms.
2. Ma e ial and me hods
2.1. Cons uc ion o ecombinan E. coli isola es exp essing
ep esen a i e β-lac amases unde low and high pe meabili y
condi ions
We designed a lib a y o up o 70 isogenic E. coli ecom-
binan isola es exp essing a ep esen a i e a ay o Amble ´
s
class A ( blaGES-1
, blaGES-5
, blaGES-7
, blaGES-15
, blaGES-20
, blaCTX-M-9
,
blaCTX-M-15
, blaSHV-12
, blaPER-1
, blaVEB-1
, blaKPC-2
, blaKPC-35
,
blaKPC-2 N132S
, blaKPC-2 L169A
, blaKPC-3
, blaKPC-31
, blaKPC-3 L167R
,
blaKPC-3 D179N
), class B ( blaVIM-1
, blaVIM-2
, blaVIM-20
, blaIMP-13
,
blaIMP-28
, blaNDM-1
, blaNDM-5
, blaNDM-7
, blaNDM-23
), class C ( blaCMY-2
,
blaDHA-1
and blaFOX-4
) and class D ( blaOXA-2
, blaOXA-15
, blaOXA-10
,
blaOXA-14
, blaOXA-48
) β-lac amases. To his end a copy o each
β-lac amase was amplified om ep esen a i e clinical isola es,
liga ed o he mul icopy plasmid pUCP-24 (gen amicin esis ance
gene casse e) and elec opo a ed in pa allel in o he E. coli TG1
and in o he OmpC- and OmpF-deficien E. coli HB4 s ains.
T ans o man s we e selec ed on LB aga pla es con aining 10
mg/L-gen amicin. Recombinan plasmids we e cha ac e ized by
MIC de e mina ion ollowing he me hodology desc ibed below.
Finally, o p ecisely de e mine he ole o double ca bapene-
mase p oduc ion, blaVIM-1
, blaIMP-28
, and blaOXA-48
ca bapenemase
encoding genes we e also amplified wi h specific p ime s and
cloned in o he mul i-copy plasmid pBGS-18 (kanamycin esis-
ance gene casse e). These ecombinan pBGS-18 plasmids we e
hen elec opo a ed in o p e iously cons uc ed E. coli TG1 and E.
coli HB4 ans o man s exp essing blaKPC-3
, blaNDM-1
o blaOXA-48
in he pUCP-24 plasmid and selec ed on LB aga pla es con ain-
ing 10 mg/L-gen amicin and 50 mg/L-kanamycin. All o he esul -
ing ans o man s we e checked by PCR o confi m he p esence o
plasmids.
2.2. An imic obial suscep ibili y es ing
Minimum inhibi o y concen a ions (MICs) o ce azidime, ce -
azidime/a ibac am, az eonam, az eonam/a ibac am, ce epime,
ce epime/enme azobac am, ce epime/ anibo bac am, ce epime/
zidebac am, zidebac am, cefide ocol, imipenem, imipenem/
elebac am, me openem, me openem/ abo bac am, me openem/
nacubac am, nacubac am, and me openem/xe ubo bac am we e
de e mined o all ans o man s in iplica e expe imen s by e e -
ence b o h mic odilu ion assays. The MICs we e de e mined using
2
T. Blanco-Ma ín, I. Alonso-Ga cía, L. González-Pin o e al. In e na ional Jou nal o An imic obial Agen s 63 (2024) 107150
ca ion-adjus ed Müelle -Hin on (MH) b o h in all cases, wi h he
excep ion o cefide ocol, which was assessed using i on-deple ed
ca ion-adjus ed MH b o h p epa ed acco ding o CLSI M100 guide-
lines [13] . Tazobac am, a ibac am, and anibo bac am we e es ed
a a fixed concen a ion o 4 mg/L, whe eas enme azobac am,
abo bac am and xe ubo bac am we e es ed a 8 mg/L. Zidebac-
am and nacubac am we e es ed a a 1:1 a io wi h ce epime
and me openem, espec i ely. EUCAST 13.1 clinical b eakpoin s
and guidelines ( h p://www.eucas .o g/clinical_b eakpoin s/ ) we e
used o e e ence pu poses. The En e obac e ales EUCAST clinical
b eakpoin o he β-lac am pa ne was used o define suscep ibil-
i y o esis ance o as ye unapp o ed combina ions. The e e ence
s ain E. coli ATCC 25922 was used as a con ol.
3. Resul s and discussion
3.1. Role o β-lac amases
Cumula i e MIC da a o he whole collec ion and each hos
s ain a e shown in Table S1, and de ailed compa a i e da a o
E. coli TG1 ans o man s p oducing single β-lac amases a e p e-
sen ed in Table 1 . The addi ion o a ibac am o ce azidime, eco -
e ed he ac i i y o ce azidime agains mos o Amble ´
s class A,
C and D β-lac amases, indica ing he high po ency o he fi s -in-
class β-lac am/DBO combina ion. Conce ning inc eases in he MIC
o ce azidime/a ibac am we e no ed o ans o man s p oducing
KPC-35 (a KPC-2 L169P mu an ; MIC = 8 mg/L) o KPC-31 (a KPC-
3 D179Y mu an ; MIC = 16 mg/L). The high e olu iona y po en ial
o KPC enzymes owa d ce azidime/a ibac am- esis ance has been
ex ensi ely epo ed since he combina ion became a ailable o
clinical use [ 14 , 15 ]. Al hough he final MIC alues we e s ill be-
low he suscep ibili y b eakpoin , impo an inc eases in MIC we e
also no ed o ans o man s p oducing PER-1 (MIC = 2 mg/L) o he
class D ex ended-spec um OXAs: OXA-14 (MIC = 4 mg/L) and OXA-
15 (MIC = 2 mg/L), a phenomenon p e iously obse ed wi h iso-
genic s ains o in pa ien s ecei ing ce azidime/a ibac am ea -
men [16] . Fo una ely, bo h PER and he ex ended-spec um OXAs
a e s ill o mino conce n in En e obac e ales since hey a e mainly
encoun e ed in P. ae uginosa . As expec ed, a ibac am pe o med
well agains all class C a ian s included, bu did no enhance
he ac i i y o ce azidime agains any o he MBL-p oducing E.
coli TG1 s ains. Az eonam/a ibac am yielded e y simila MICs o
hose o ce azidime/a ibac am agains mos o class A, C and D
β-lac amase ans o man s. The mos in e es ing ea u e o hese
new combina ion is p obably i s ou s anding ac i i y agains MBL
p oduce s, which in all cases exhibi ed an az eonam/a ibac am
MIC o ≤0.06 mg/L. The findings clea ly indica e ha his combina-
ion would be an addi ion o upmos impo ance o ea in ec ions
caused by MBL-p oducing s ains. Howe e , az eonam/a ibac am-
esis an lineages o NDM-5 p oducing E. coli ha e al eady been
de ec ed due o he p esence o amino-acid inse ions in PBP-3 (-
YRIN-, - YRIP- o -YRIK-; in ol ed in dec eased az eonam affini y)
o he p oduc ion o ce ain AmpC a ian s wi h inc eased ac i i y
owa d az eonam (e.g., CMY-42) [ 17 , 18 ].
Analysis o he po ency o ce epime in combina ion wi h he
β-lac amase inhibi o s enme azobac am, anibo bac am and zide-
bac am also e ealed p omising esul s. Ce epime/enme azobac am
yielded MICs in he suscep ibili y ange agains mos o class A,
C and D β-lac amases (MICs ≤0.06-0.5 mg/L) bu was no ac i e
agains MBLs [19] . The combina ion has been shown o esis he
ac i i y o AmpCs and ESBLs when challenged agains la ge col-
lec ions o clinical isola es and is also ac i e agains some p o-
duce s o KPC o OXA-48 enzymes ( he la e o which is almos
always accompanied by CTX-M a ian s) [20] . The po en ial use
o his combina ion as a ca bapenem-spa ing egimen aimed a
a o ding ca bapenems looking o a educ ion in he bu den o
ca bapenem- esis an o ganism in he hospi al en i onmen is a
niche ha dese es explo a ion. Simila ly, ce epime/ anibo bac am
also demons a ed e y s ong po ency agains all class A, C and
D β-lac amases (MICs ≤0.06 mg/L), bu also p o ided good p o-
ec ion o ce epime agains some MBL p oduce s. The combina-
ion was ac i e agains VIM- ype enzymes (MICs ≤0.06-1 mg/L).
Howe e , as p e iously shown IMP- ype enzymes we e no inhib-
i ed, as indica ed by he negligible e ec on he MICs o ce epime
o IMP-28 o IMP-13 (MICs = 4–8 mg/L o bo h ce epime and ce-
epime/ anibo bac am) [21] . Tanibo bac am also e ec i ely educed
he MIC o ce epime o NDM p oduce s, al hough bo de line sus-
cep ibili y was no ed o some a ian s (e.g., NDM-5; MIC = 8 mg/L),
p obably explained by he high MICs o ce epime ha could no
be e ec i ely dec eased o below he suscep ibili y b eakpoin .
Ce epime/zidebac am exhibi ed he b oades ange o he apeu ic
co e age agains he whole se o E. coli TG1 ans o man s, which
in all cases yielded MICs anging om ≤0.03 o 0.25 mg/L. The
po ency obse ed o ce epime/zidebac am seems o esul om
he syne gis ically imp o ed bac e ial killing by ce epime and zide-
bac am, as highligh ed by he high in insic ac i i y o zidebac-
am agains all he ans o man s ega dless o he β-lac amase
p oduced (MICs = 0.06–0.5 mg/L) [22] . To da e, esis ance o ce-
epime/zidebac am has a ely been de ec ed in clinical isola es,
al hough specific mu a ions a ec ing PBP-2 ha e been de ec ed
in esis an mu an s ob ained in i o [23] . Al oge he , hese e-
sul s suppo he p omising ole ha ce epime/ anibo bac am and
ce epime/zidebac am may ha e agains ca bapenemase-p oducing
En e obac e ales, pa icula ly hose ca ying MBLs.
Cefide ocol was highly ac i e agains he whole collec ion o
E. coli TG1 ans o man s, wi h MICs anging om ≤0.06 o 2
mg/L. Cefide ocol ac i i y was significan ly a ec ed by p oduc-
ion o SHV-12, PER-1 and NDM-like enzymes, which as p e i-
ously epo ed inc eased he MICs o he cephalospo in o 1, 1
and 2 mg/L, espec i ely. Howe e , in acco dance wi h ou da a,
inc eased cefide ocol MICs and clinical esis ance a e pe sis en ly
epo ed o s ains o En e obac e ales ca ying NDM ca bapene-
mases, hus a guing o close moni o iza ion o i s ac i i y agains
MBL p oduce s. Ca bapenems we e es ed in combina ion wi h
elebac am, in he case o imipenem, o pa ne ed wi h abo bac-
am, nacubac am o xe ubo bac am, in he case o me openem.
Due o high baseline suscep ibili y o bo h ca bapenems in he
E. coli TG1 hos , imipenem/ elebac am, me openem/ abo bac am,
me openem/xe ubo bac am and me openem/nacubac am we e, as
expec ed, ac i e agains all ans o man s ca ying class A, C
and D β-lac amases, wi h MICs anging in all cases be ween
0.03 and 0.25 mg/L. In his ega d, he highes MICs o hese
ca bapenem/ β-lac amase inhibi o combina ions co esponded o
he ans o man ca ying he OXA-48 ca bapenemase, agains
which he addi ion o none o hese inhibi o s educed he MIC
o he β-lac am pa ne . P oduc ion o class B ca bapenemases
(and pa icula ly NDM) was associa ed wi h g ea e esis ance o
hese ca bapenem combina ions. Xe ubo bac am o nacubac am
dec eased 4 wo- old dilu ions he MIC o me openem agains
NDM enzymes, highligh ing hei p omising ole agains MBLs in
he nea u u e.
3.2. In e play be ween β-lac amases and dec eased ou e memb ane
pe meabili y
Analysis o he MIC da a o he 35 OmpC/OmpF-deficien E. coli
HB4 ans o man s allowed us o ob ain a de ailed iew o how de-
c eased β-lac am in e naliza ion could ampli y le els o esis ance
o he agen s es ed ( he global cumula i e e ec is ep esen ed
in Table S1, and de ailed MICs a e shown in Table 2 ). Compa ed
wi h E. coli TG1, clinical esis ance o ce azidime/a ibac am in E.
coli HB4 ex ended o PER-1 (MIC = 2–16 mg/L), OXA-15 (MIC = 2–
3
T. Blanco-Ma ín, I. Alonso-Ga cía, L. González-Pin o e al. In e na ional Jou nal o An imic obial Agen s 63 (2024) 107150
Table 1
An ibio ic suscep ibili y da a o newly de eloped
β-lac ams and β-lac am/ β-lac amase inhibi o combina ions agains isogenic E. coli TG1 ans o man s exp essing he mos ele an class A, B, C and D β-lac amases ound in
En e obac e ales.
S ain Amble
class
Main Pheno ype MIC (mg/L)
a
CAZ
(R > 4)
C/A
(R > 8)
ATM
(R > 4)
A/A
(R > 4)
FEP
(R > 4)
F/T
(R > 4)
F/E
(R > 4)
F/Z
(R > 4)
ZID
(NA)
FDC
(R > 2)
IMP
(R > 4)
I/R
(R > 2)
MEM
(R > 8)
M/V
(R > 8)
M/X
(R > 8)
M/N
(R > 8)
NAC
(NA)
E. coli TG1 - Wild- ype 0.125 0.125 ≤0.06 ≤0.06 ≤0.06 ≤0.06 ≤0.06 ≤0.03 0.06 ≤0.06 0.25 0.125 ≤0.06 ≤0.06 ≤0.06 ≤0.03 2
GES-1
A
ESBL 4 0.125 ≤0.06 ≤0.06 0.125 ≤0.06 ≤0.06 ≤0.03 0.125 ≤0.06 0.125 0.125 ≤0.06 ≤0.06 ≤0.06 ≤0.03 2
GES-5 Ca bapenemase 0.25 0.125 ≤0.06 ≤0.06 ≤0.06 ≤0.06 ≤0.06 ≤0.03 0.125 ≤0.06 0.25 0.125 ≤0.06 ≤0.06 ≤0.06 ≤0.03 2
GES-7 ESBL 32 0.25 0.5 ≤0.06 0.125 ≤0.06 ≤0.06 0.06 0.25 ≤0.06 0.25 0.25 ≤0.06 ≤0.06 ≤0.06 ≤0.03 2
GES-15 ESBL 8 0.125 0.125 ≤0.06 0.125 ≤0.06 ≤0.06 0.06 0.25 ≤0.06 0.125 0.125 ≤0.06 ≤0.06 ≤0.06 ≤0.03 2
GES-20 Ca bapenemase 0.25 0.125 ≤0.06 ≤0.06 ≤0.06 ≤0.06 ≤0.06 ≤0.03 0.25 ≤0.06 0.25 0.25 ≤0.06 ≤0.06 ≤0.06 ≤0.03 2
CTX-M-9 ESBL 0.5 0.125 2 ≤0.06 1 ≤0.06 ≤0.06 0.125 0.125 ≤0.06 0.125 0.125 ≤0.06 ≤0.06 ≤0.06 ≤0.03 2
CTX-M-15 ESBL 32 0.125 32 ≤0.06 32 ≤0.06 ≤0.06 0.125 0.125 ≤0.06 0.25 0.125 ≤0.06 ≤0.06 ≤0.06 ≤0.03 2
SHV-12 ESBL > 64 0.25 > 64 0.125 4 ≤0.06 ≤0.06 0.125 0.125 1 0.25 0.125 ≤0.06 ≤0.06 ≤0.06 ≤0.03 2
PER-1 ESBL > 64 2 > 64 2 8 ≤0.06 ≤0.06 0.125 0.125 1 0.25 0.125 ≤0.06 ≤0.06 ≤0.06 ≤0.03 2
VEB-1 ESBL > 64 0.25 32 ≤0.06 1 ≤0.06 ≤0.06 0.06 0.125 0.25 0.125 0.125 ≤0.06 ≤0.06 ≤0.06 ≤0.03 2
KPC-2 Ca bapenemase 16 0.25 > 64 0.125 16 ≤0.06 ≤0.06 0.125 0.125 ≤0.06 1 0.25 0.5 ≤0.06 ≤0.06 0.125 2
KPC-35 (KPC-2 L169P) ESBL > 64 8 1 ≤0.06 1 ≤0.06 ≤0.06 0.125 0.125 ≤0.06 0.25 0.125 ≤0.06 ≤0.06 ≤0.06 ≤0.03 2
KPC-2 N132S Ca bapenemase ≤0.06 ≤0.06 ≤0.06 ≤0.06 ≤0.06 ≤0.06 ≤0.06 ≤0.03 0.125 ≤0.06 0.25 0.25 0.125 ≤0.06 ≤0.06 0.06 2
KPC-2 L169A ESBL 16 1 8 ≤0.06 8 ≤0.06 ≤0.06 0.125 0.125 ≤0.06 1 0.25 0.125 ≤0.06 ≤0.06 0.06 2
KPC-3 Ca bapenemase 32 0.25 64 ≤0.06 4 ≤0.06 ≤0.06 0.125 0.125 ≤0.06 1 0.125 0.25 ≤0.06 ≤0.06 0.06 2
KPC-31 (KPC-3 D179Y) ESBL 64 16 0.5 ≤0.06 1 ≤0.06 ≤0.06 0.125 0.125 0.125 0.125 0.125 ≤0.06 ≤0.06 ≤0.06 ≤0.03 2
KPC-3 L167R Ca bapenemase 4 0.125 4 ≤0.06 0.25 ≤0.06 ≤0.06 0.125 0.125 ≤0.06 0.25 0.125 ≤0.06 ≤0.06 ≤0.06 ≤0.03 2
KPC-3 D179N ESBL 32 8 1 0.125 1 ≤0.06 ≤0.06 0.125 0.125 ≤0.06 0.25 ≤0.06 ≤0.06 ≤0.06 ≤0.06 ≤0.03 2
VIM-1
B
Ca bapenemase > 64 > 64 ≤0.06 ≤0.06 32 1 32 0.125 0.125 0.125 2 2 0.5 0.5 0.5 0.5 2
VIM-2 Ca bapenemase 1 1 ≤0.06 ≤0.06 ≤0.06 ≤0.06 ≤0.06 ≤0.03 0.06 ≤0.06 0.25 0.25 ≤0.06 ≤0.06 ≤0.06 ≤0.03 2
VIM-20 Ca bapenemase 2 2 ≤0.06 ≤0.06 0.25 ≤0.06 0.25 0.125 0.125 ≤0.06 0.25 0.25 ≤0.06 ≤0.06 ≤0.06 0.06 2
IMP-13 Ca bapenemase > 64 > 64 ≤0.06 ≤0.06 8 8 4 0.125 0.125 ≤0.06 0.25 0.25 0.125 0.125 0.125 0.125 2
IMP-28 Ca bapenemase > 64 > 64 ≤0.06 ≤0.06 8 4 8 0.125 0.125 ≤0.06 0.25 0.25 ≤0.06 ≤0.06 ≤0.06 0.06 2
NDM-1 Ca bapenemase > 64 > 64 ≤0.06 ≤0.06 32 1 16 0.125 0.125 1 2 1 2 2 0.5 0.5 2
NDM-5 Ca bapenemase > 64 > 64 ≤0.06 ≤0.06 > 64 8 > 64 0.125 0.25 2 32 32 16 16 1 1 2
NDM-7 Ca bapenemase > 64 > 64 ≤0.06 ≤0.06 > 64 4 > 64 0.125 0.125 2 16 16 8 8 1 1 2
NDM-23 Ca bapenemase > 64 > 64 ≤0.06 ≤0.06 32 1 32 0.125 0.125 2 4 2 2 2 0.5 0.5 2
CMY-2
C
Ex ended-spec um cephamycinase > 64 0.5 16 0.25 1 ≤0.06 0.5 0.25 0.5 0.125 0.5 0.25 ≤0.06 ≤0.06 ≤0.06 ≤0.03 2
DHA-1 Ex ended-spec um cephamycinase 32 0.125 4 ≤0.06 0.125 ≤0.06 ≤0.06 ≤0.03 0.25 ≤0.06 0.125 0.125 ≤0.06 ≤0.06 ≤0.06 ≤0.03 2
FOX-4 Ex ended-spec um cephamycinase > 64 1 2 ≤0.06 0.25 ≤0.06 0.125 0.06 0.25 ≤0.06 0.25 0.25 ≤0.06 ≤0.06 ≤0.06 ≤0.03 2
OXA-2
D
Na ow-spec um oxacillinase 2 ≤0.06 ≤0.06 ≤0.06 ≤0.06 ≤0.06 ≤0.06 0.06 0.25 0.125 0.25 0.125 ≤0.06 ≤0.06 ≤0.06 ≤0.03 2
OXA-15 ESBL 32 2 ≤0.06 ≤0.06 0.25 ≤0.06 ≤0.06 0.125 0.125 0.25 0.25 0.125 ≤0.06 ≤0.06 ≤0.06 ≤0.03 2
OXA-10 Na ow-spec um oxacillinase 0.125 ≤0.06 0.5 0.125 0.25 ≤0.06 ≤0.06 0.125 0.125 ≤0.06 0.25 0.125 ≤0.06 ≤0.06 ≤0.06 ≤0.03 2
OXA-14 ESBL 32 4 0.5 0.125 0.5 ≤0.06 0.125 0.125 0.125 0.25 0.25 0.125 ≤0.06 ≤0.06 ≤0.06 ≤0.03 2
OXA-48 Ca bapenemase 0.25 ≤0.06 ≤0.06 ≤0.06 0.125 ≤0.06 ≤0.06 0.06 0.125 ≤0.06 0.5 0.25 0.25 0.25 ≤0.06 0.25 2
CAZ: ce azidime; C/A: ce azidime/a ibac am; ATM: az eonam; A/A: az eonam/a ibac am; FEP: ce epime; F/E: ce epime/enme azobac am; F/T: ce epime/ anibo bac am; F/Z: ce epime/zidebac am; ZID: zidebac am; FDC: ce-
fide ocol; IMP: imipenem; IMP/REL: imipenem/ elebac am; MEM: me openem; M/V: me openem/ abo bac am; M/X: me openem/xe ubo bac am; M/N: me openem/nacubac am; NAC: nacubac am; NA: No a ailable
a EUCAST b eakpoin s indica ed o En e obac e ales.
4
T. Blanco-Ma ín, I. Alonso-Ga cía, L. González-Pin o e al. In e na ional Jou nal o An imic obial Agen s 63 (2024) 107150
Table 2
An ibio ic suscep ibili y da a o newly de eloped
β-lac ams and β-lac am/ β-lac amase inhibi o combina ions agains isogenic E. coli HB4 ans o man s exp essing he mos ele an class A, B, C and D β-lac amases ound in
En e obac e ales.
S ain Amble class Main Pheno ype MIC (mg/L)
a
CAZ
(R > 4)
C/A
(R > 8)
ATM
(R > 4)
A/A
(R > 4)
FEP
(R > 4)
F/T
(R > 4)
F/E
(R > 4)
F/Z
(R > 4)
ZID
(NA)
FDC
(R > 2)
IMP
(R > 4)
I/R
(R > 2)
MEM
(R > 8)
M/V
(R > 8)
M/X
(R > 8)
M/N
(R > 8)
NAC
(NA)
E. coli HB4 - OmpC/OmpF po in-deficien 1 0.5 0.25 0.125 0.5 0.5 0.5 0.25 0.25 ≤0.06 0.25 0.125 ≤0.06 ≤0.06 ≤0.06 ≤0.03 2
GES-1
A
ESBL 16 1 1 0.125 2 0.5 0.5 0.25 0.5 ≤0.06 0.125 0.125 0.25 0.125 ≤0.06 0.125 2
GES-5 Ca bapenemase 2 0.5 0.25 0.125 0.5 0.5 0.5 0.25 0.25 ≤0.06 0.5 0.125 4 0.25 ≤0.06 0.5 2
GES-7 ESBL > 64 1 4 0.125 4 0.5 0.5 0.25 0.5 ≤0.06 0.125 0.125 0.125 ≤0.06 ≤0.06 0.125 2
GES-15 ESBL > 64 2 1 0.125 8 0.5 0.5 0.5 0.5 ≤0.06 0.125 0.125 0.25 0.125 ≤0.06 0.125 2
GES-20 Ca bapenemase 2 0.5 0.25 0.125 1 0.5 0.5 0.25 0.5 ≤0.06 1 0.125 2 0.125 ≤0.06 0.5 2
CTX-M-9 ESBL 4 0.5 16 0.125 64 0.5 0.5 2 2 ≤0.06 0.25 0.125 0.25 0.125 ≤0.06 0.125 2
CTX-M-15 ESBL > 64 1 > 64 0.125 > 64 1 0.5 1 2 0.125 0.25 0.125 0.25 0.125 0.125 0.125 2
SHV-12 ESBL > 64 4 > 64 0.5 > 64 1 64 1 1 2 0.25 0.125 1 1 0.25 0.25 2
PER-1 ESBL > 64 16 > 64 16 > 64 1 64 1 1 2 0.25 ≤0.06 0.5 0.25 0.125 0.25 2
VEB-1 ESBL > 64 2 > 64 1 64 0.5 0.5 0.25 0.5 0.5 0.125 ≤0.06 0.125 0.125 0.125 0.125 2
KPC-2 Ca bapenemase > 64 2 > 64 1 > 64 1 > 64 4 4 0.125 64 0.5 > 64 0.5 ≤0.06 2 2
KPC-35 (KPC-2 L169P) ESBL > 64 32 8 0.25 64 2 1 1 1 1 0.25 0.125 2 0.5 0.125 1 2
KPC-2 N132S Ca bapenemase 1 0.5 0.25 0.125 0.5 0.5 0.5 0.125 0.25 ≤0.06 4 4 16 0.5 0.125 1 2
KPC-2 L169A ESBL > 64 16 32 0.25 64 2 2 2 4 0.125 4 0.25 32 0.5 0.125 1 2
KPC-3 Ca bapenemase > 64 4 > 64 0.125 > 64 1 > 64 1 1 0.25 16 0.125 > 64 0.5 0.125 1 2
KPC-31 (KPC-3 D179Y) ESBL > 64 64 4 0.25 64 2 0.5 0.5 0.5 1 0.25 0.125 1 0.5 0.125 1 2
KPC-3 L167R Ca bapenemase 32 1 32 0.125 16 0.5 2 0.25 0.5 ≤0.06 4 1 4 0.25 0.125 1 2
KPC-3 D179N ESBL > 64 16 8 0.25 8 0.5 0.25 0.125 0.125 0.25 0.25 0.125 0.125 ≤0.06 ≤0.06 0.125 2
VIM-1
B
Ca bapenemase > 64 > 64 0.25 0.125 64 8 64 0.125 0.5 0.125 2 2 2 2 2 1 2
VIM-2 Ca bapenemase 8 8 0.25 0.125 1 0.5 1 0.25 0.25 ≤0.06 0.25 0.25 1 1 0.5 0.5 2
VIM-20 Ca bapenemase 16 16 0.25 0.125 4 0.5 4 0.125 0.25 ≤0.06 1 1 2 2 0.5 1 2
IMP-13 Ca bapenemase > 64 > 64 0.25 0.125 32 32 32 0.125 0.25 ≤0.06 1 1 8 8 8 1 2
IMP-28 Ca bapenemase > 64 > 64 0.25 0.125 32 32 32 0.125 0.125 ≤0.06 1 1 16 16 16 2 2
NDM-1 Ca bapenemase > 64 > 64 0.25 0.25 > 64 16 > 64 0.125 0.125 2 16 16 32 8 16 1 2
NDM-5 Ca bapenemase > 64 > 64 0.25 0.25 > 64 > 64 > 64 0.25 0.25 2 > 64 > 64 > 64 > 64 64 4 2
NDM-7 Ca bapenemase > 64 > 64 0.25 0.25 > 64 > 64 > 64 0.25 0.25 4 > 64 > 64 > 64 > 64 64 4 2
NDM-23 Ca bapenemase > 64 > 64 0.25 0.25 > 64 32 > 64 0.125 0.25 2 8 8 64 32 8 2 2
CMY-2
C
Ex ended-spec um
cephamycinase
> 64 2 64 1 32 0.5 8 2 4 0.25 1 0.25 1 0.25 ≤0.06 0.25 2
DHA-1 Ex ended-spec um
cephamycinase
> 64 1 32 0.25 2 0.5 2 1 4 ≤0.06 0.5 0.125 1 0.25 ≤0.06 0.25 2
FOX-4 Ex ended-spec um
cephamycinase
> 64 4 8 0.25 4 0.5 4 0.5 1 ≤0.06 0.25 0.125 0.5 0.5 0.125 0.25 2
OXA-2
D
Na ow-spec um
oxacillinase
16 1 0.25 0.125 1 0.5 0.5 0.25 0.5 0.25 0.5 0.5 2 2 0.125 1 2
OXA-15 ESBL 64 16 2 0.125 4 1 2 0.5 0.5 0.5 0.125 0.125 0.5 0.25 0.25 0.25 2
OXA-10 Na ow-spec um
oxacillinase
2 0.5 8 1 8 0.5 4 0.25 0.5 ≤0.06 0.25 0.25 4 1 0.125 2 2
OXA-14 ESBL > 64 64 16 1 16 1 4 0.5 0.5 0.5 0.25 0.125 1 0.25 0.5 0.5 2
OXA-48 Ca bapenemase 1 0.5 0.25 0.25 2 0.5 2 0.25 0.25 ≤0.06 2 1 8 8 ≤0.06 2 2
CAZ: ce azidime; C/A: ce azidime/a ibac am; ATM: az eonam; A/A: az eonam/a ibac am; FEP: ce epime; F/E: ce epime/enme azobac am; F/T: ce epime/ anibo bac am; F/Z: ce epime/zidebac am; ZID: zidebac am; FDC: ce-
fide ocol; IMP: imipenem; I/R: imipenem/ elebac am; MEM: me openem; M/V: me openem/ abo bac am; M/X: me openem/xe ubo bac am; M/N: me openem/nacubac am; NAC: nacubac am; NA: No a ailable
a EUCAST b eakpoin s indica ed o En e obac e ales .
5

T. Blanco-Ma ín, I. Alonso-Ga cía, L. González-Pin o e al. In e na ional Jou nal o An imic obial Agen s 63 (2024) 107150
16 mg/L) and OXA-14 (MIC = 4– 64 mg/L). Mo eo e , some en-
zymes such as SHV-12, KPC-3 o FOX-4 inc eased esis ance o
le els close o he suscep ibili y b eakpoin (MICs = 4 mg/L). The
e ec o PER-1 in his backg ound was also amplified o az e-
onam/a ibac am (MIC = 16 mg/L), al hough in gene al was poo ly
a ec ed by he loss o po ins (MIC = 0.125–1 mg/L o he es o
isola es). Ce epime was one o he subs a es agains which lesions
in po ins con e ed g ea e de ec s on i s ac i i y. The combina-
ion wi h anibo bac am was unca ed agains MBLs, as he MIC
inc eased o 8 mg/L o highe in all cases excep VIM-2 and VIM-
20. Ce epime/enme azobac am showed dec eased ac i i y agains
se e al key class A a ge s, such as SHV-12 (MIC = 64 mg/L), PER-1
(MIC = 64 mg/L), KPC-2 (MIC > 64 mg/L) and KPC-3 (MIC > 64 mg/L),
bu also bo de line MICs (4–8 mg/L) o se e al class C and class D
a ian s. The e ec o low pe meabili y was no as e iden o ce-
epime/zidebac am, which main ained wi h 100% ac i i y, p obably
esul ing om he mechanis ic syne gy igge ed by PBP-3 a ge -
ing o ce epime wi h he enhancing e ec o zidebac am.
Cefide ocol main ained a es o ac i i y simila o hose ob-
se ed in E. coli TG1, highligh ing ha in e naliza ion h ough
side opho es can bypass any pe meabili y de ec . The MIC o ca -
bapenem combina ions expe imen ed a majo inc ease agains he
HB4 collec ion, a ac ha helped o cla i y he p o ec i e e ec s
(i any) o he di e en inhibi o s. Imipenem/ elebac am was ac i e
agains mos o he class A and C enzymes and class D a ian s
wi h ESBL ac i i y. While imipenem/ elebac am has shown o se-
lec inac i a ing mu a ions in OmpK36 when used o comba KPC
in ec ions, i was able o educe he MIC o imipenem by be ween
5 and 7 wo- old dilu ions agains he E. coli HB4 de i a i es p o-
ducing KPC-2 (MIC = 0.5 mg/L) o KPC-3 (MIC = 0.125 mg/L) [24] .
In e es ingly, he only class A β-lac amase able o con e clinical
esis ance o imipenem/ elebac am was he KPC-2 N132S a ian
(MIC = 4 mg/L). O no e, he eplacemen o aspa agine by se ine a
posi ion 132, loca ed on he YSN iad o he KPC-2 enzyme, has
ecen ly been desc ibed in i o and demons a ed o con e de-
c eased elebac am suscep ibili y in e ms o IC50
, Ki app
and k2
/ K
[25] . Vabo bac am po en ia ed he ac i i y o me openem agains
all class A p oduce s, including key a ge s such as KPC-2 (MIC = 0.5
mg/L) and KPC-3 (MIC = 0.5 mg/L), bu was inac i e agains class
B (MICs = 1- > 64 mg/L) enzymes o OXA-48 (MIC = 8 mg/L). I s ac-
i i y was also significan ly educed by OXA-2 (MIC = 2 mg/L) and
OXA-10 enzymes (MIC = 1 mg/L), which significan ly inc ease he
MIC o ca bapenems unde condi ions o low pe meabili y [26] .
Me openem/xe ubo bac am was he me openem-based combina-
ion wi h highes ac i i y se ine- ype enzymes, pa icula ly ac i e
agains KPC-2 (MIC = ≤0.06 mg/L), KPC-3 (MIC = 0.125 mg/L) and
OXA-48 (MIC ≤0.06 mg/L). Howe e , he p e iously obse ed an i-
MBL efficacy was no e iden (MICs = 0.5–64 mg/L), a ac p oba-
bly explained by he p esence o non- unc ional po ins which ha e
been shown o a ec he in e naliza ion o xe ubo bac am [ 27 , 28 ].
Finally, me openem/nacubac am demons a ed globally highe MIC
alues han o he compounds bu i s ac i i y ex ended o he
whole collec ion.
3.3. E ec s o double ca bapenemase p oduc ion
We finally cons uc ed double ans o man s o es ima e he
po en ial he apeu ic ole o hese inno a i e op ions agains
he eme ging h ea o double ca bapenemase p oduc ion
( Table 3 ). Double ca bapenemase p oduc ion usually esul ed
in he loss o ce azidime/a ibac am, imipenem/ elebac am,
me openem/ abo bac am, ce epime/enme azobac am and ce-
epime/ anibo bac am, mos ly due o p oduc ion o class B
enzymes (pa icula ly no iceable in he E. coli HB4 s ain). In
his ega d, p obably he mos challenging associa ions o ca -
bapenemases we e hose esul ing om he combina ion o
Table 3
An ibio ic suscep ibili y da a o E. coli TG1 and E. coli HB4 ecombinan isola es exp essing double ca bapenemases.
S ain Amble
class
Main Pheno ype MIC (mg/L)
a
CAZ
(R > 4)
C/A
(R > 8)
ATM
(R > 4)
A/A
(R > 4)
FEP
(R > 4)
F/T
(R > 4)
F/E
(R > 4)
F/Z
(R > 4)
ZID
(NA)
FDC
(R > 2)
IMP
(R > 4)
I/R
(R > 2)
MEM
(R > 8)
M/V
(R > 8)
M/X
(R > 8)
M/N
(R > 8)
NAC
(NA)
E. coli TG1 - Wild- ype 0.125 0.125 ≤0.06 ≤0.06 ≤0.06 ≤0.06 ≤0.06 ≤0.03 0.06 ≤0.06 0.25 0.125 ≤0.06 ≤0.06 ≤0.03 ≤0.06 2
KPC-3 + VIM-1 A + B
Double ca bapenemase 64 32 64 ≤0.06 16 0.25 8 0.25 0.25 ≤0.06 2 0.5 1 0.125 0.125 0.125 2
KPC-3 + IMP-28 Double ca bapenemase 64 64 64 ≤0.06 4 2 2 0.125 0.125 ≤0.06 8 0.5 2 0.25 0.25 0.5 2
KPC-3 + OXA-48 A + D Double ca bapenemase 16 ≤0.25 32 ≤0.06 4 ≤0.25 ≤0.25 0.125 0.125 ≤0.06 2 0.25 2 0.125 0.125 ≤0.06 2
NDM-1 + OXA-48
B + D
Double ca bapenemase > 256 > 256 ≤0.06 ≤0.06 64 8 64 0.06 0.06 2 8 8 8 4 1 0.5 2
OXA-48 + VIM-1 Double ca bapenemase 128 32 ≤0.06 ≤0.06 8 0.25 8 0.25 0.125 ≤0.06 1 0.5 0.25 0.125 0.25 ≤0.06 2
OXA-48 + IMP-28 Double ca bapenemase 64 32 ≤0.06 ≤0.06 4 4 4 0.06 0.06 ≤0.06 1 1 0.5 0.25 0.125 ≤0.06 2
E. coli HB4 - OmpC/OmpF po in-deficien 1 0.5 0.25 0.125 0.5 0.5 0.5 0.25 0.25 ≤0.06 0.25 0.125 ≤0.06 ≤0.06 ≤0.06 ≤0.03 2
KPC-3 + VIM-1 A + B
Double ca bapenemase > 256 128 > 64 0.25 128 8 128 1 1 0.125 8 1 32 4 4 1 2
KPC-3 + IMP-28 Double ca bapenemase 256 128 > 64 0.25 128 16 16 0.5 1 ≤0.06 8 0.5 32 8 8 1 2
KPC-3 + OXA-48 A + D Double ca bapenemase > 256 0.5 > 64 0.25 256 32 64 0.5 0.5 ≤0.06 8 0.5 16 8 0.5 1 2
NDM-1 + OXA-48
B + D
Double ca bapenemase > 256 > 256 0.25 0.25 > 256 32 256 0.25 0.125 2 4 4 32 32 8 1 2
OXA-48 + VIM-1 Double ca bapenemase > 256 128 0.5 0.25 128 4 128 0.25 0.25 0.125 4 2 8 8 1 1 2
OXA-48 + IMP-28 Double ca bapenemase 256 128 0.5 0.25 32 32 32 0.25 0.125 ≤0.06 2 1 8 8 16 1 2
CAZ: ce azidime; C/A: ce azidime/a ibac am; ATM: az eonam; A/A: az eonam/a ibac am; FEP: ce epime; F/E: ce epime/enme azobac am; F/T: ce epime/ anibo bac am; F/Z: ce epime/zidebac am; ZID: zidebac am; FDC: ce-
fide ocol; IMP: imipenem; I/R: imipenem/ elebac am; MEM: me openem; M/V: me openem/ abo bac am; M/X: me openem/xe ubo bac am; M/N: me openem/nacubac am; NAC: nacubac am; NA: No a ailable
a EUCAST b eakpoin s indica ed o En e obac e ales .
6
T. Blanco-Ma ín, I. Alonso-Ga cía, L. González-Pin o e al. In e na ional Jou nal o An imic obial Agen s 63 (2024) 107150
KPC-3 + IMP-28 and NDM-1 + OXA-48, which espec i ely educed
he ac i i y o me openem/xe ubo bac am in E. coli HB4 o
con e ed educed cefide ocol suscep ibili y. Hope ully, az e-
onam/a ibac am (MIC ≤0.06–0.25 mg/L), ce epime/zidebac am
(MIC = 0.06–1 mg/L), cefide ocol (MIC ≤0.06–2 mg/L) and o a lesse
ex en me openem/nacubac am (MIC ≤0.06–1 mg/L) will ep esen
aluable op ions agains isola es showing such combina ions o
esis ance mechanisms.
4. Conclusions
The collec ion he e e alua ed only includes ecombinan iso-
la es o E. coli and hus hei implica ions o o he species should
be p ecisely de e mined in u u e s udies. Howe e , ou findings
highligh he p omising s abili y o hese new β-lac ams and β-
lac am/ β-lac amase inhibi o combina ions agains he main β-
lac amases ci cula ing in En e obac e ales, including when hey a e
combined wi h low pe meabili y o addi ional ca bapenemases. Al-
hough some o he op ions e alua ed he e appea o mi iga e he
u gency o new compounds able o esis he ac ion o MBL o
some ex en , d ug de elopmen e o s and su eillance a e needed
o comba he sp ead o some enzymes, such as NDM, which can
esis he ac i i y o las gene a ion inhibi o s o limi he ac i i y
o cefide ocol.
Decla a ions
Funding: This wo k was suppo ed by he Ins i u o de Salud Ca -
los III (ISCIII, p ojec s PI22/01212 , PI20/01212 and PI21/00704 )and
co- unded by he Eu opean Union. This wo k was also suppo ed
by Me ck Sha p & Dohme (MSD) h ough he In es iga o Ini i-
a ed S udies P og am. The esea ch was also unded by Cen o
de In es igación Biomédica en Red de En e medades In ecciosas
(CIBERINFEC, CB21/13/0 0 055 , CB21/13/0 0 099 , CB21/13/0 0 095 and
CB21/13/0 0 012 ), he Spanish Ne wo k o Resea ch in In ec ious
Diseases (REIPI, N °RD16/0016/0004 and N °RD16/0016/0006 ), in-
eg a ed in he Na ional Plan o Scien ific Resea ch, De elop-
men and Technological Inno a ion 2013–2016 and unded by he
ISCIII-Gene al Subdi ec ion o Assessmen and P omo ion o he
Resea ch-Eu opean Regional De elopmen Fund (FEDER) “A way
o making Eu ope.” The s udy was also unded by he Axen-
cia Galega de Inno ación (GAIN), Conselle ía de Inno ación, Con-
selle ía de Emp ego e Indus ia ( IN607D2021/12 o A.B. and IN607A
2016/22 o G.B.). This esea ch was also suppo ed by Pe sonal-
ized and p ecision medicine g an om he Ins i u o de Salud Ca -
los III (MePRAM P ojec , PMP22/0 0 092 ), Ins i u o de Salud Ca -
los III, Minis e io de Ciencia e Inno ación. T.B.-M. was financially
suppo ed by he ISCIII p ojec PI20/00686 and by he Rio Ho -
ega p og am (ISCIII, CM23/0 0 095 ). I.A.-G. was financially suppo ed
by he Rio Ho ega p og am (ISCIII, CM21/0 0 076 ) and by he Juan
Rodés p og am (ISCIII, JR23/0 0 036 ). L.G.-P. was financially sup-
po ed by he ISCIII p ojec PI21/00704 and by he PFIS p o-
g am (ISCIII, FI23/0 0 074 ). J.C.V.-U. was financially suppo ed by he
Xun a de Galicia ( IN606B-2022/009 ). J.A.-S. was financially sup-
po ed by he Juan Rodés p og am (ISCIII, JR21/0 0 026 ).
Compe ing in e es s: Me ck Sha p & Dohme p o ided elebac am
powde and cefide ocol was p o ided by Shionoghi. Comme cial
supplie s did no exe cise any con ol o e he conduc o epo -
ing o he esea ch.
J.C.V.-U. has ecei ed hono a ia o lec u es and/o p esen a-
ions om MSD. A.O. has ecei ed g an s o con ac s om MSD,
Wockha d and Shionogi, and consul ing ees and hono a ia o
lec u es and/o p esen a ions om MSD, Pfize and Shionogi. G.B.
has ecei ed unding and s udy ma e ials om MSD, g an s con-
ac s om MSD, Pfize , ABAC The apeu ics and Roche, consul ing
ees and hono a ia o lec u es and/o p esen a ions om MSD, Sh-
ionogi Pfize , Roche and Mena ini, and suppo o a ending mee -
ings and/o a els om Pfize . J.A.-S. has ecei ed hono a ia o
lec u es and/o p esen a ions om Shionogi and Ad anz Pha ma.
All o he au ho s: none o decla e.
E hical app o al: No equi ed.
Sequence in o ma ion: No applicable.
Supplemen a y ma e ials
Supplemen a y ma e ial associa ed wi h his a icle can be
ound, in he online e sion, a doi:10.1016/j.ijan imicag.2024.
107150 .
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8