research article fecal colonization with extended-spectrum...

Research ArticleFecal Colonization with Extended-Spectrum Beta-Lactamase andAmpC-Producing Escherichia coli

Mohamed H Al-Agamy12 Taghrid S El Mahdy3 and Atef M Shibl14

1Department of Pharmaceutics Microbiology Division College of Pharmacy King Saud University PO Box 2457Riyadh 11451 Saudi Arabia2Department of Microbiology and Immunology Faculty of Pharmacy Al-Azhar University Cairo Egypt3Microbiology and Immunology Department Faculty of Pharmacy Helwan University Cairo Egypt4College of Medicine Alfaisal University Riyadh Saudi Arabia

Correspondence should be addressed to Mohamed H Al-Agamy elagamy71yahoocom

Received 18 December 2015 Accepted 10 May 2016

Academic Editor Wejdene Mansour

Copyright copy 2016 Mohamed H Al-Agamy et alThis is an open access article distributed under the Creative Commons AttributionLicense which permits unrestricted use distribution and reproduction in anymedium provided the originalwork is properly cited

Background Extended-spectrum120573-lactamases (ES120573Ls) andAmpC 120573-lactamases cause120573-lactam resistance in Escherichia coli Fecalcolonization by ES120573L- andor AmpC-positive E coli is a source of nosocomial infectionsMethods In order to investigate inpatientfecal colonization by ES120573Ls and AmpC antibiotic sensitivity tests were conducted andminimum inhibitory concentrations (MICs)were determined using the disk diffusion method and E-test respectively Characterization of ES120573L and AmpC was performedusing E-test strips and a set of PCRs and DNA sequence analyses were used to characterize the ES120573L and AmpC genes ResultsThe whole collection of E coli isolates (119899 = 50) was sensitive to imipenem tigecycline colistin and fosfomycin while 26 of theisolates showed reduced susceptibility to ceftazidime (MIC ge 4 120583gmL) ES120573L was phenotypically identified in 26 (1350) of caseswhile AmpC activity was detected in two ES120573L-producing E coli isolates All ES120573L-producing E coli were positive for the CTX-M gene eleven isolates carried 119887119897119886CTX-M-15 and two isolates carried 119887119897119886CTX-M-14 gene Two CTX-M-positive E coli isolates carried119887119897119886CMY-2 Conclusions The alimentary tract is a significant reservoir for ES120573L- andor AmpC-producing E coli which may lead tonosocomial infection

1 Introduction

A remarkable increase in fecal colonization rates with ex-tended-spectrum beta-lactamase- (ES120573L-) AmpC-plasmidmediated- andor carbapenemases-producing Enterobac-teriaceae has been reported in many regions worldwide[1 2] Infections caused by Enterobacteriaceae which areresistant to 120573-lactams are coupled with the inappropriateuse of antibiotics andor a prolonged period of hospitaladmission The rising use of carbapenems for empiricaltreatment of nosocomial infections has led to fast global dis-semination of carbapenemase-positive enterobacterial strains[3] ES120573Ls arise through point mutations in TEM-1TEM-2 and SHV-1 However over the last three decades non-TEM and non-SHV ES120573Ls strains have been detected pri-marily CTX-M Enterobacteriaceae that produce CTX-Menzymes have shown rapid and concerning dissemination

and have been documented as the most prevalent etio-logical infectious agents [4] ES120573L confers resistance topenicillins cephalosporins and monobactam (aztreonam)but they are susceptible to cephamycins (cefoxitin andcefotetan) and carbapenems (imipenem meropenem anddoripenem) and are typically reserved by inhibitors ofAmbler class A 120573-lactamase (clavulanic acid tazobactamor sulbactam) Most ES120573Ls can hydrolyze fourth-generationcephalosporins While AmpC 120573-lactamases confer resistanceto penicillins third-generation cephalosporins monobac-tam and cephamycins they are sensitive to carbapenem andare not inhibited by 120573-lactamase inhibitors however they areinhibited by cloxacillin [5ndash7]

Numerous studies in Saudi Arabia have focused on theidentification of ES120573L-producing strains from clinical speci-mens [4] but there are few reports on the fecal colonizationof ES120573L-producing isolates in Saudi Arabia Therefore in

Hindawi Publishing CorporationBioMed Research InternationalVolume 2016 Article ID 3704150 7 pageshttpdxdoiorg10115520163704150

2 BioMed Research International

the present study we determine the incidence of ES120573L-andor AmpC cephalosporinase-producing Escherichia coliisolates in human fecal flora and investigated the genesencoding the corresponding enzymes

2 Materials and Methods

21 Bacterial Identification Fifty different E coli isolates wereisolated from 50 stool samples of different inpatients carriersunder nonoutbreak conditions at a hospital in Riyadh SaudiArabia from April 2014 to June 2014 Briefly fresh stoolspecimens were aseptically collected and transported to themicrobiology laboratory Stool samples were suspended insterile phosphate-buffered saline pH 7 A 100120583L volume wasdirectly inoculated onto blood agar andEosinMethyleneBlueagar (Oxoid Microbiology Products Hampshire UK) After48 h incubation at 37∘C the isolated organisms were identi-fied by conventional procedures and automated identificationsystems with the API20E identification kit (bioMerieuxMarcy lrsquoEtoile France)These isolates were preserved in brainheart infusion broth containing 20 glycerol at minus70∘C

22 Phenotypic Detection of ES120573L The isolates showingreduced susceptibility to ceftazidime (CAZ) cefotaxime(CTX) or aztreonam (ATM) (minimum inhibitory concen-tration (MIC) ge 1 120583gmL or zone diameter le 22mm) wereselected for screening of ES120573L production (Clinical and Lab-oratory Standards Institute (CLSI) 2014) E-test ES120573L stripswere used in accordance with the manufacturerrsquos instruc-tions to evaluate ES120573L production The CAZceftazidime+ clavulanate- (CAZCAL-) ES120573L E-test strip was used todetect ES120573L production The test is considered positive ifthe ratio of MIC of CAZCAL is ge8 To inhibit AmpC120573-lactamase the CAZCAL-ES120573L E-test was carried outon cloxacillin Mueller-Hinton agar and the results wereinterpreted in a similar manner

23 Phenotypic Detection of AmpC The isolates showingreduced susceptibility to cefoxitin (FOX) or cefotetan (CTT)(zone diameter of 18 or 16mm resp) were selected forscreening of AmpC enzyme production [9] The phenotypicdetection test consists of a strip containing CTT on one endand CTT-cloxacillin (CTTCXT) on the other end Ratios oftheMICs of CTTCXT ge 8 are considered to indicate positiveAmpC 120573-lactamase production

24 Susceptibility Testing MICs for the isolates showing aphenotype of producing ES120573L and AmpC activities weredetermined by using E-test strips (bioMerieux MarcylrsquoEtoile France) Interpretation was based on the Clinical andLaboratory Standards Institute (CLSI) criteria [9]Escherichiacoli ATCC 25922 strains were used as reference strainsThe following antibiotics were tested piperacillin (PIP)piperacillintazobactam (TZP) CAZ CAZCAL CTXcefepime (FEP) ATM FOX CTT CTTCXT imipenem(IMI) gentamicin (GM) amikacin (AK) ciprofloxacin (CI)colistin (COL) tigecycline (TGC) and fosfomycin (FOS)

25 Screening for the Presence of 120573-Lactamase Genes Theisolate was cultured in 2mL of Tryptic Soy Broth (DifcoFranklin Lakes NJ USA) A 200120583L volume of overnightculture was heated at 99∘C in a heat block for 10minThe obtained DNA was used in polymerase chain reaction(PCR) assays on a Techne FlexigeneThermal Cycler (TechneDuxford Cambridge UK) Positive and negative controlswere included in all PCR assays All PCR products wereanalyzed on 08 agarose gels (incorporated with 05mgLethidium bromide) and then visualized under UV light(Pharmacia LKB Biotechnology AB Gothenburg Sweden)and photographed using a documentation system (CE DP-CF-011C European Union)

The PCR primers used are listed in Table 1 The primerswere used to search for class A 120573-lactamase genes (119887119897119886TEM119887119897119886SHV 119887119897119886OXA-1 and 119887119897119886CTX-M families) and class C 120573-lactamase genes (119887119897119886CMY 119887119897119886MOX 119887119897119886FOX 119887119897119886DHA 119887119897119886ACC119887119897119886ACT 119887119897119886MIR 119887119897119886EBC 119887119897119886CIT and 119887119897119886BIL) PCR assays wereconducted as previously described [8]

26 Sequencing of 120573-Lactamase Genes Purification of PCRamplicons was performed using a PCR purification kit(Qiagen Hilden Germany) PCR products of bla genes weresequenced on both strands using PCR primers to determinetheir molecular types DNA sequences were analyzed usingthe ABI 3100 Genetic Analyzer (Applied Biosystems FosterCity CA USA) according to the manufacturerrsquos recommen-dations

3 Results

31 Bacterial Identification Fifty fecal E coli samples wereisolated randomly from hospitalized patients in RiyadhSaudi Arabia The patients were treated for noninfectiousdiseases under nonoutbreak conditions Escherichia coli iso-lates were identified manually and according to the API20Eidentification kit (bioMerieux Marcy lrsquoEtoile France)

32 Characterization of 119887119897119886ES120573L and 119887119897119886AmpC Thirteen of the50 E coli isolates which showed reduced susceptibility toCAZ CTX or ATM (MIC ge 1 120583gmL or inhibition zonele 22mm) were selected for screening of ES120573L and AmpCenzyme production using CAZCAL-ES120573L and CTTCXT-AmpC E-test strips Thirteen isolates were positive for ES120573Land two ES120573L-positive isolates producedAmpC120573-lactamase

PCRwas used to detect 119887119897119886ES120573L genes andAmpC plasmid-mediated genes in ES120573L- and AmpC-positive E coli isolates(119899 = 13) The results of PCR and DNA sequencing of blagenes are shown in Table 2

Eleven of 13 ES120573L-producing E coli isolates were found tocontain CTX-M-15 while two isolates harbored 119887119897119886CTX-M-14CMY-2-positive isolates (119899 = 2) were concomitant withCTX-M-15 All ES120573L-producing E coli isolates (119899 = 13)were positive for 119887119897119886TEM-1 while eight (615) isolates carried119887119897119886OXA-1 In contrast three (23) isolates were found tocontain 119887119897119886SHV-1

BioMed Research International 3

Table1Prim

ersu

sedfora

mplificatio

nof

thetested120573-la

ctam

aseg

enes

(Dallenn

eetal2010

[8])

PCRtype

Target

Prim

erSequ

ence

ofprim

ers(51015840-31015840

)Amplified

prod

ucts

(bp)

Multip

lexIT

EMSHV

and

OXA-

1-like

TEM

MultiT

SO-T

for

CATT

TCCG

TGTC

GCC

CTTA

TTC

800

MultiT

SO-T

rev

CGTT

CATC

CATA

GTT

GCC

TGAC

SHV

MultiT

SO-S

for

AGCC

GCT

TGAG

CAAAT

TAAAC

713

MultiT

SO-S

rev

ATCC

CGCA

GAT

AAAT

CACC

AC

OXA-

1MultiT

SO-O

for

GGCA

CCAG

ATTC

AAC

TTTC

AAG

564

MultiT

SO-O

rev

GAC

CCCA

AGTT

TCCT

GTA

AGTG

Multip

lexIICT

X-M

grou

p1grou

p2andgrou

p9

CTX-

Mgrou

p1

MultiC

TXMGp1

for

TTAG

GAART

GTG

CCGCT

GYA

688

MultiC

TXMGp1-2

rev

CGAT

ATCG

TTGGTG

GTR

CCAT

CTX-

Mgrou

p2

MultiC

TXMGp2

for

CGTT

AAC

GGCA

CGAT

GAC

404

MultiC

TXMGp1-2

rev

CGAT

ATCG

TTGGTG

GTR

CCAT

CTX-

Mgrou

p9

MultiC

TXMGp9

for

TCAAG

CCTG

CCGAT

CTGGT

561

MultiC

TXMGp9

rev

TGAT

TCTC

GCC

GCT

GAAG

CTX-

Mgrou

p825

CTX-

Mgrou

p825

CTX-

Mg825for

AAC

RCRC

AGAC

GCT

CTAC

326

CTX-

Mg825rev

TCGAG

CCGGAASG

TGTY

AT

Multip

lexIIIA

CCFOX

MOX

DHAC

ITand

EBC

ACC-

1and

ACC-

2MultiC

aseA

CCfor

CACC

TCCA

GCG

ACTT

GTT

AC346

MultiC

aseA

CCrev

GTT

AGCC

AGCA

TCAC

GAT

CC

FOX-

1toFO

X-5

MultiC

aseFOX

for

CTAC

AGTG

CGGGTG

GTT

T162

MultiC

aseFOX

rev

CTAT

TTGCG

GCC

AGGTG

AMOX-

1MOX-

2CM

Y-1CM

Y-8to

CMY-11

andCM

Y-19

MultiC

aseM

OX

for

GCA

ACAAC

GAC

AAT

CCAT

CCT

895

MultiC

aseM

OX

rev

GGGAT

AGGCG

TAAC

TCTC

CCAA

DHA-

1and

DHA-

2MultiC

aseD

HA

for

TGAT

GGCA

CAGCA

GGAT

ATTC

997

MultiC

aseD

HA

rev

GCT

TTGAC

TCTT

TCGGTA

TTCG

LAT-1toLA

T-3BIL-1CM

Y-2to

CMY-7

CMY-12

toCM

Y-18and

CMY-21

toCM

Y-23

MultiC

aseC

ITfor

CGAAG

AGGCA

ATGAC

CAGAC

538

MultiC

aseC

ITrev

ACGGAC

AGGGTT

AGGAT

AGY

ACT-1and

MIR-1

MultiC

aseEBC

for

CGGTA

AAG

CCGAT

GTT

GCG

683


Table2Antim

icrobialsusceptib

ilityprofi

leso

fES120573

L-andAmpC

-produ

cing

Ecoliiso

latesfrom

fecalsam

ples

andassociated

resistancep

atterns

Isolates

number

119864-te

stMIC

(mgL)

Resistanceg

enes

PIP

TZP

CTX

CAZ

CAZCA

LFE

PAT

MFO

XCT

TCT

TCX

TIM

IGM

AK

CICO

LTG

CFO

SEC

1gt256lt1gt256

32322

48

0032

025

lt05lt05

006

05

21lt0016

025

0016

TEM-1+C

TX-M

-15+

OXA-

1EC

2gt256lt1gt256

161605

416

0065

025

lt05lt05

012

153

025lt0016

025lt0016

TEM-1+C

TX-M

-15+

+SHV-1

EC3

gt256

64gt256gt256

gt324

12gt256

0125

025

lt05lt05

025

643

4lt0016

0125lt0016

TEM-1+C

TX-M

-15+

OXA-

1EC

4gt256

8gt256

32320064

816

0032

006

lt05lt05

006

124

4lt0016

0125lt0016

TEM-1+C

TX-M

-15+

OXA-

1EC

5gt256

32gt256

161605

412

0125

0125lt05lt05

003

83

6lt0016

025lt0016

TEM-1+C

TX-M

-15+

SHV-1

EC6

gt256

8gt256gt256

gt324

192gt256

025

025

lt05lt05

006

192

48gt32lt0016

025lt0016

TEM-1+C

TX-M

-15+

OXA-

1EC

7gt256

8gt256gt256

gt324

128

192

025

025

lt05lt05

012

128

16gt32lt0016

025lt0016

TEM-1+C

TX-M

-15+

OXA-

1EC

8gt256lt1gt256gt256

gt324

96128

025

025

lt05lt05

006

9605

025lt0016

025

0125

TEM-1+C

TX-M

-14+

OXA-

1EC

9gt256

16gt256

44006

43

30032

0032lt05lt05

006

82

025lt0016

038lt0016

TEM-1+C

TX-M

-15

EC10

gt256

4gt256

660125

46

0032

0032lt05lt05

0125

161

05lt0016

025lt0016

TEM-1+C

TX-M

-15+

SHV-1

EC11

gt256lt1gt256

48gt321

824

0032

0032lt05lt05

025

232

1lt0016

025lt0016

TEM-1+C

TX-M

-14+

OXA-

1EC

12gt256gt256gt256gt256gt32gt4

48gt256

6432

3232

025gt256

192

075lt0016

025

0016

TEM-1+C

TX-M

-15+

CMY-2+

OXA-

1EC

13gt256

128gt256gt256gt32gt4

64gt256

6448

gt32gt32

025

128

4gt32lt0016

038

0016

TEM-1+C

TX-M

-15+

CMY-2

PIP

piperacillin

TZP

piperacillintazobactam

CAZ

cefta

zidime

CAZCA

Lcefta

zidimecefta

zidime+cla

vulanicacidC

TXc

efotaxim

eFE

Pcefepime

ATMa

ztreon

amF

OX

cefoxitin

CTT

cefotetan

CTTCX

Tcefotetancefotetan+clo

xacillin

IMIim

ipenem

GMgentamicinA

Kam

ikacinC

Iciprofl

oxacinC

OLcolistin

TGC

tigecyclin

eFO

Sfosfo

mycin


33 Antimicrobial Resistance Pattern of ES120573L and AmpCEnzyme-Positive Isolates The MICs of 17 antimicrobialagents were determined for E coli fecal isolates The resultsof the susceptibility pattern for ES120573L and AmpC enzyme-producing E coli isolates are illustrated in Table 2

4 Discussion

The human and animal alimentary tracts are vital reservoirsfor ES120573L- carbapenemases- and AmpC enzyme-producingEnterobacteriaceae Patient-to-patient transmission of resis-tant microorganisms may occur in hospitals [10 11] Theoveruse of antibiotics has recently been associated with theemergence of resistant intestinal bacteria particularly ES120573L-carbapenemases- and AmpC enzyme-producing Enterobac-teriaceae Numerous studies have demonstrated that expo-sure to 120573-lactam antibiotics is a risk factor for the selection ofmultidrug-resistant E coli [12] Therefore the current studyexamined the antimicrobial resistance patterns of fecal Ecoli isolates as well as the molecular basis for their 120573-lactamresistance mechanisms using phenotypic and genotypicmethods The present study included 50 patients admitted toa hospital in Riyadh Saudi Arabia from April 2014 to June2014 These patients were treated for noninfectious diseasesunder nonoutbreak conditions Fifty fecal stool specimenscollected from the 50 patients were cultured on blood agarand EMB agar as described in Materials and Methods Fiftysuspected E coli isolates were selected and the isolates wereidentified by conventional procedures and using the API20Eidentification kit All isolates were identified as E coli

Production of 120573-lactamases is the main mechanism of120573-lactam resistance in Gram-negative bacteria including Ecoli [6 7] Several 120573-lactamases (ES120573Ls AmpC enzymes andcarbapenemases) have been previously reported in fecal Ecoli isolates [13ndash15]

In the present study 100 of 50 E coli isolates were foundto be sensitive to imipenem and 13 (26) of 50 isolates wereresistant or showed reduced susceptibility to CAZ CTX-M or ATM The carbapenem susceptibility results indicatedthat our isolates did not harbor carbapenemase while 26(1350) of the E coli isolates harbored ES120573L andor AmpC 120573-lactamase Two of 13 isolates exhibited reduced susceptibilityto cephamycins (FOX and CTT)

Phenotypic screening for the presence of different typesof 120573-lactamases was conducted using E coli isolatesThirteenE coli isolates were selected for screening of ES120573L andAmpC 120573-lactamase production using CAZCAL-ES120573L andCTTCXT-AmpC E-test strips were used to detect ES120573Lproduction Using phenotypic detection methods all isolateswere found to produce ES120573L while two isolates phenotyp-ically produced AmpC enzyme A battery of PCR assayswas conducted to detect 119887119897119886ES120573L genes and AmpC plasmid-mediated genes in the 13 E coli isolates Therefore class Aand class C 120573-lactamase genes were tested The PCR-purifiedproduct was subjected to DNA sequencing to identify thegene variants The results of molecular characterization ofbla genes are shown in Table 2 PCR amplification andDNA sequencing analyses of the PCR products showed

that all isolates possessed a CTX-M-type ES120573L and that119887119897119886CTX-M-15 was present in 1 isolate while two isolates con-tained 119887119897119886CTX-M-14 Other CTX-M families were not detectedThe gene encoding CMY-2 enzyme was detected in two Ecoli isolates CMY-2-positive isolates are concomitant withCTX-M-15 All E coli isolates (119899 = 13) were positivefor 119887119897119886TEM-1 while 615 and 23 of the isolates contained119887119897119886OXA-1 and 119887119897119886SHV-1 respectivelyThe increase in expressionof the AmpC 120573-lactamases may mask the recognition ofES120573Ls [16] Therefore in the present study the genotypicmethods revealed that all 13 strains were ES120573L CTX-M-positive while phenotypic methods showed that 11 strainswere ES120573L-positive and two strains were AmpC enzyme-positive AmpC-producing strains producingCTX-M-15mayact as a dormant reservoir for ES120573Ls

Numerous studies have documented a remarkableincrease in intestinal colonization rates with ES120573L- andAmpC enzyme-producing Enterobacteriaceae in manycountries [2 10 11 14ndash17]The prevalence of ES120573L-producingE coli fecal isolates varies widely from country to countryfrom region to region and at different time periods Ahigh incidence of fecal carriage rate of ES120573L-producing Ecoli has been observed in Asia Africa and South America[13 14 18ndash21] while a significantly lower prevalence ofES120573L-producing E coli fecal isolates was reported in mostEuropean countries [22 23] In Argentina the rate of fecalcarriage of Enterobacteriaceae-resistant strains to third-generation cephalosporins was 268 [20] Villar et al [20]reported that 2022 and 67 of fecal strains were colonizedby ES120573L- and AmpC enzyme-producing Enterobacteriaceae[20] In Egypt Al-Agamy et al reported that 226 and322 of hospitalized patients were colonized by ES120573L-and AmpC-positive E coli respectively The 119887119897119886CTX-M-likegene was the predominant ES120573L gene detected in 714of ES120573L-producing E coli isolates [21] In a recent studyin Egypt Bassyouni et al reported that 21 and 3 ofpatients were colonized by ES120573L- and AmpC-producing Ecoli respectively They also found that 119887119897119886SHV gene was thepredominant ES120573L gene detected in 818 of the resistant Ecoli isolates [13] In Korea 203 of fecal Enterobacteriaceaemembers were ES120573Ls [19] In India the prevalence ofES120573L-positive E coli isolates was 19 in healthy volunteersfrom the community [14] In Libya 134 and 67 of E coliisolates were ES120573Ls- and AmpC-positive respectively [18]In a previous study in Saudi Arabia 177 of strains werefound to be ES120573L-positive [24] A high (261) prevalencewas detected in inpatients followed by outpatients (154)and the lowest prevalence rate (131) was detected in healthyindividuals [24] In the present study the prevalence of ES120573L-producing E coli was 26 Despite differences in the dateand region of isolation the prevalence of fecal carriage rateof ES120573L in the present study (26) was in agreement withthe prevalence (261) reported by Kader et al In contrastthe prevalence rate of ES120573L-producing Enterobacteriaceaewas 29 among healthy Swedish children Escherichia colicontaining CTX-M 120573-lactamase predominated and onlyone E coli isolate harbored genes encoding for CMY [22]The carriage rate of ES120573L- and AmpC enzyme-producing Ecoli was 357 and 238 among 84 Danish army recruits


respectively 119887119897119886CTX-M-14 gene was the predominant ES120573Lgene detected in three (100) ES120573L-producing E coliisolates while 119887119897119886CMY-2 was detected in two AmpC enzyme-producing E coli isolates [23] In Spain the prevalence ofES120573L and AmpC enzyme carriers was 506 and 059respectively [25] 119887119897119886CTX-M genes were the ES120573L dominatinggenes (9615) and CTX-M-14 was the most prevalent gene(50) followed by CTX-M-15 (40) CMY-2 was the mostprevalent gene (8125) followed by DHA-1 (1875) [25]

MICs were determined for ES120573L- and AmpC enzyme-producing E coli (119899 = 13) isolates The results of MIC areshown in Table 2

In conclusion a high incidence of carriage of ES120573L-positive E coli fecal isolates among hospitalized patients inRiyadh was detected reaching 26 with 119887119897119886CTX-M-15 (846)being the most predominant gene The emergence of fecalcarriage of CMY-2-producing E coli among hospitalizedpatients has been reported to be 4These outcomes empha-size the importance of the intestinal tract as a reservoir forES120573L- and AmpC enzyme-producing E coli which may leadto nosocomial infection The admission of colonized fecalcarriers of ES120573L- and AmpC-positive E coli to the medicalsetting increases the possibility of other patients acquiringinfection in the same hospital Our results emphasize thenecessity for continuous surveillance in hospitals to detectthe ES120573L- AmpC enzyme- and carbapenemase-producingstrains andmultidrug strains as well applying effective strate-gies for antimicrobial therapy and infection control measuresto decrease the abuse and misuse of antimicrobial agentsagainst resistant strains and to prevent their spread

Competing Interests

The authors declare that they have no competing interests

Acknowledgments

The authors extend their appreciation to the Deanship ofScientific Research at King Saud University for funding thework through the research group Project no RGP-038

References

[1] A Bhargava K Hayakawa E Silverman et al ldquoRisk factors forcolonization due to carbapenem-resistant enterobacteriaceaeamong patients exposed to long-term acute care and acute carefacilitiesrdquo Infection Control and Hospital Epidemiology vol 35no 4 pp 398ndash405 2014

[2] P-L Woerther C Burdet E Chachaty and A AndremontldquoTrends in human fecal carriage of extended-spectrum 120573-lactamases in the community toward the globalization of CTX-Mrdquo Clinical Microbiology Reviews vol 26 no 4 pp 744ndash7582013

[3] N Karah B Haldorsen N O Hermansen et al ldquoEmergenceof OXA-carbapenemase- and 16S rRNA methylase-producinginternational clones of Acinetobacter baumannii in NorwayrdquoJournal of Medical Microbiology vol 60 no 4 pp 515ndash521 2011

[4] S Yezli A M Shibl and Z A Memish ldquoThemolecular basis of120573-lactamase production in Gram-negative bacteria from Saudi

Arabiardquo Journal of Medical Microbiology vol 64 no 2 pp 127ndash136 2015

[5] M Guzman-Blanco J A Labarca M V Villegas and EGotuzzo ldquoExtended spectrum 120573-lactamase producers amongnosocomial Enterobacteriaceae in Latin Americardquo BrazilianJournal of Infectious Diseases vol 18 no 4 pp 421ndash433 2014

[6] Y Pfeifer A Cullik andWWitte ldquoResistance to cephalosporinsand carbapenems in Gram-negative bacterial pathogensrdquo Inter-national Journal ofMedicalMicrobiology vol 300 no 6 pp 371ndash379 2010

[7] R Canton and T M Coque ldquoThe CTX-M beta-lactamasepandemicrdquo Current Opinion in Microbiology vol 9 no 5 pp466ndash475 2006

[8] C Dallenne A da Costa D Decre C Favier and G ArletldquoDevelopment of a set of multiplex PCR assays for the detectionof genes encoding important 120573-lactamases in Enterobacteri-aceaerdquo Journal of Antimicrobial Chemotherapy vol 65 no 3 pp490ndash495 2010

[9] Clinical and Laboratory Standards Institute (CLSI) ldquoPer-formance standards for antimicrobial susceptibility testingTwenty-fourth informational supplement performance stan-dards for antimicrobial susceptibility testingrdquoDocumentM100-24 Clinical and Laboratory Standards Institute (CLSI) WaynePa USA 2014

[10] J A Severin E S Lestari W Kloezen et al ldquoFaecal carriageof extended-spectrum 120573-lactamase-producing Enterobacteri-aceae among humans in Java Indonesia in 2001-2002rdquo TropicalMedicine and International Health vol 17 no 4 pp 455ndash4612012

[11] B Li J-Y Sun Q-Z Liu L-Z Han X-H Huang and Y-X Ni ldquoHigh prevalence of CTX-M 120573-lactamases in faecalEscherichia coli strains from healthy humans in Fuzhou ChinardquoScandinavian Journal of Infectious Diseases vol 43 no 3 pp170ndash174 2011

[12] G V Sanchez R N Master J A Karlowsky and J M BordonldquoIn vitro antimicrobial resistance of urinary Escherichia coliisolates among US outpatients from 2000 to 2010rdquo Antimicro-bial Agents andChemotherapy vol 56 no 4 pp 2181ndash2183 2012

[13] R H Bassyouni S N Gaber and A AWegdan ldquoFecal carriageof extended-spectrum 120573-lactamase- and AmpC- producingEscherichia coli among healthcare workersrdquo Journal of Infectionin Developing Countries vol 9 no 3 pp 304ndash308 2015

[14] D Mathai V A Kumar B Paul et al ldquoFecal carriage ratesof extended-spectrum 120573-lactamase-producing Escherichia coliamong antibiotic naive healthy human volunteersrdquo MicrobialDrug Resistance vol 21 no 1 pp 59ndash64 2015

[15] S B Joslashrgensen Oslash Samuelsen A Sundsfjord et al ldquoHighprevalence of faecal carriage of ESBL-producing Enterobacteri-aceae in Norwegian patients with gastroenteritisrdquo ScandinavianJournal of Infectious Diseases vol 46 no 6 pp 462ndash465 2014

[16] N O Yilmaz N Agus E Bozcal O Oner and A UzelldquoDetection of plasmid-mediated AmpC 120573-lactamase in E coliand K pneumoniaerdquo Indian Journal of Medical Microbiologyvol 31 no 1 pp 53ndash59 2013

[17] T M H Bui I Hirai S Ueda et al ldquoCarriage of Escherichiacoli producing CTX-M-type extended-spectrum 120573-lactamasein healthy Vietnamese individualsrdquo Antimicrobial Agents andChemotherapy vol 59 no 10 pp 6611ndash6614 2015

[18] S F Ahmed M M M Ali Z K Mohamed T A Moussa andJ D Klena ldquoFecal carriage of extended-spectrum 120573-lactamasesand AmpC-producing Escherichia coli in a Libyan communityrdquo


Annals of Clinical Microbiology and Antimicrobials vol 13article 22 2014

[19] Y J Ko H-W Moon M Hur C-M Park S E Cho andY-M Yun ldquoFecal carriage of extended-spectrum 120573-lactamase-producing Enterobacteriaceae in Korean community and hos-pital settingsrdquo Infection vol 41 no 1 pp 9ndash13 2013

[20] H E Villar M N Baserni and M B Jugo ldquoFaecal carriage ofESBL-producing enterobacteriaceae and carbapenem-resistantGram-negative bacilli in community settingsrdquo Journal of Infec-tion in Developing Countries vol 7 no 8 pp 630ndash634 2013

[21] M H Al-Agamy M S Ali M M Salem and T R MohamedldquoFaecal colonization by extended-spectrum beta-lactamase-producing Escherichia coli from hospitalized patientsrdquo NewEgyptian Journal of Microbiology vol 19 pp 285ndash314 2008

[22] J Kaarme Y Molin B Olsen and A Melhus ldquoPrevalence ofextended-spectrum beta-lactamase-producing Enterobacteri-aceae in healthy Swedish preschool childrenrdquo Acta Paediatricavol 102 no 6 pp 655ndash660 2013

[23] A M Hammerum C H Lester L Jakobsen and L J PorsboldquoFaecal carriage of extended-spectrum 120573-lactamase-producingand AmpC 120573-lactamase-producing bacteria among Danisharmy recruitsrdquo Clinical Microbiology and Infection vol 17 no4 pp 566ndash568 2011

[24] A A Kader A Kumar and K A Kamath ldquoFecal car-riage of extended-spectrum120573-lactamase-producingEscherichiacoli and Klebsiella pneumoniae in patients and asymptomatichealthy individualsrdquo Infection Control and Hospital Epidemiol-ogy vol 28 no 9 pp 1114ndash1116 2007

[25] A Garrido C Seral M J Gude et al ldquoCharacterization ofplasmid-mediated 120573-lactamases in fecal colonizing patients inthe hospital and community setting in Spainrdquo Microbial DrugResistance vol 20 no 4 pp 301ndash304 2014

Submit your manuscripts athttpwwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Anatomy Research International

PeptidesInternational Journal of


Hindawi Publishing Corporation httpwwwhindawicom

International Journal of

Volume 2014

Zoology


Molecular Biology International

GenomicsInternational Journal of


The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014


BioinformaticsAdvances in

Marine BiologyJournal of



Signal TransductionJournal of


BioMed Research International

Evolutionary BiologyInternational Journal of



Biochemistry Research International

ArchaeaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014


Genetics Research International


Advances in

Virolog y

Hindawi Publishing Corporationhttpwwwhindawicom

Nucleic AcidsJournal of

Volume 2014

Stem CellsInternational



Enzyme Research



Microbiology


the present study we determine the incidence of ES120573L-andor AmpC cephalosporinase-producing Escherichia coliisolates in human fecal flora and investigated the genesencoding the corresponding enzymes

2 Materials and Methods

21 Bacterial Identification Fifty different E coli isolates wereisolated from 50 stool samples of different inpatients carriersunder nonoutbreak conditions at a hospital in Riyadh SaudiArabia from April 2014 to June 2014 Briefly fresh stoolspecimens were aseptically collected and transported to themicrobiology laboratory Stool samples were suspended insterile phosphate-buffered saline pH 7 A 100120583L volume wasdirectly inoculated onto blood agar andEosinMethyleneBlueagar (Oxoid Microbiology Products Hampshire UK) After48 h incubation at 37∘C the isolated organisms were identi-fied by conventional procedures and automated identificationsystems with the API20E identification kit (bioMerieuxMarcy lrsquoEtoile France)These isolates were preserved in brainheart infusion broth containing 20 glycerol at minus70∘C

22 Phenotypic Detection of ES120573L The isolates showingreduced susceptibility to ceftazidime (CAZ) cefotaxime(CTX) or aztreonam (ATM) (minimum inhibitory concen-tration (MIC) ge 1 120583gmL or zone diameter le 22mm) wereselected for screening of ES120573L production (Clinical and Lab-oratory Standards Institute (CLSI) 2014) E-test ES120573L stripswere used in accordance with the manufacturerrsquos instruc-tions to evaluate ES120573L production The CAZceftazidime+ clavulanate- (CAZCAL-) ES120573L E-test strip was used todetect ES120573L production The test is considered positive ifthe ratio of MIC of CAZCAL is ge8 To inhibit AmpC120573-lactamase the CAZCAL-ES120573L E-test was carried outon cloxacillin Mueller-Hinton agar and the results wereinterpreted in a similar manner

23 Phenotypic Detection of AmpC The isolates showingreduced susceptibility to cefoxitin (FOX) or cefotetan (CTT)(zone diameter of 18 or 16mm resp) were selected forscreening of AmpC enzyme production [9] The phenotypicdetection test consists of a strip containing CTT on one endand CTT-cloxacillin (CTTCXT) on the other end Ratios oftheMICs of CTTCXT ge 8 are considered to indicate positiveAmpC 120573-lactamase production

24 Susceptibility Testing MICs for the isolates showing aphenotype of producing ES120573L and AmpC activities weredetermined by using E-test strips (bioMerieux MarcylrsquoEtoile France) Interpretation was based on the Clinical andLaboratory Standards Institute (CLSI) criteria [9]Escherichiacoli ATCC 25922 strains were used as reference strainsThe following antibiotics were tested piperacillin (PIP)piperacillintazobactam (TZP) CAZ CAZCAL CTXcefepime (FEP) ATM FOX CTT CTTCXT imipenem(IMI) gentamicin (GM) amikacin (AK) ciprofloxacin (CI)colistin (COL) tigecycline (TGC) and fosfomycin (FOS)

25 Screening for the Presence of 120573-Lactamase Genes Theisolate was cultured in 2mL of Tryptic Soy Broth (DifcoFranklin Lakes NJ USA) A 200120583L volume of overnightculture was heated at 99∘C in a heat block for 10minThe obtained DNA was used in polymerase chain reaction(PCR) assays on a Techne FlexigeneThermal Cycler (TechneDuxford Cambridge UK) Positive and negative controlswere included in all PCR assays All PCR products wereanalyzed on 08 agarose gels (incorporated with 05mgLethidium bromide) and then visualized under UV light(Pharmacia LKB Biotechnology AB Gothenburg Sweden)and photographed using a documentation system (CE DP-CF-011C European Union)

The PCR primers used are listed in Table 1 The primerswere used to search for class A 120573-lactamase genes (119887119897119886TEM119887119897119886SHV 119887119897119886OXA-1 and 119887119897119886CTX-M families) and class C 120573-lactamase genes (119887119897119886CMY 119887119897119886MOX 119887119897119886FOX 119887119897119886DHA 119887119897119886ACC119887119897119886ACT 119887119897119886MIR 119887119897119886EBC 119887119897119886CIT and 119887119897119886BIL) PCR assays wereconducted as previously described [8]

26 Sequencing of 120573-Lactamase Genes Purification of PCRamplicons was performed using a PCR purification kit(Qiagen Hilden Germany) PCR products of bla genes weresequenced on both strands using PCR primers to determinetheir molecular types DNA sequences were analyzed usingthe ABI 3100 Genetic Analyzer (Applied Biosystems FosterCity CA USA) according to the manufacturerrsquos recommen-dations

3 Results

31 Bacterial Identification Fifty fecal E coli samples wereisolated randomly from hospitalized patients in RiyadhSaudi Arabia The patients were treated for noninfectiousdiseases under nonoutbreak conditions Escherichia coli iso-lates were identified manually and according to the API20Eidentification kit (bioMerieux Marcy lrsquoEtoile France)

32 Characterization of 119887119897119886ES120573L and 119887119897119886AmpC Thirteen of the50 E coli isolates which showed reduced susceptibility toCAZ CTX or ATM (MIC ge 1 120583gmL or inhibition zonele 22mm) were selected for screening of ES120573L and AmpCenzyme production using CAZCAL-ES120573L and CTTCXT-AmpC E-test strips Thirteen isolates were positive for ES120573Land two ES120573L-positive isolates producedAmpC120573-lactamase

PCRwas used to detect 119887119897119886ES120573L genes andAmpC plasmid-mediated genes in ES120573L- and AmpC-positive E coli isolates(119899 = 13) The results of PCR and DNA sequencing of blagenes are shown in Table 2

Eleven of 13 ES120573L-producing E coli isolates were found tocontain CTX-M-15 while two isolates harbored 119887119897119886CTX-M-14CMY-2-positive isolates (119899 = 2) were concomitant withCTX-M-15 All ES120573L-producing E coli isolates (119899 = 13)were positive for 119887119897119886TEM-1 while eight (615) isolates carried119887119897119886OXA-1 In contrast three (23) isolates were found tocontain 119887119897119886SHV-1


Table1Prim

ersu

sedfora

mplificatio

nof

thetested120573-la

ctam

aseg

enes

(Dallenn

eetal2010

[8])

PCRtype

Target

Prim

erSequ

ence

ofprim

ers(51015840-31015840

)Amplified

prod

ucts

(bp)

Multip

lexIT

EMSHV

and

OXA-

1-like

TEM

MultiT

SO-T

for

CATT

TCCG

TGTC

GCC

CTTA

TTC

800

MultiT

SO-T

rev

CGTT

CATC

CATA

GTT

GCC

TGAC

SHV

MultiT

SO-S

for

AGCC

GCT

TGAG

CAAAT

TAAAC

713

MultiT

SO-S

rev

ATCC

CGCA

GAT

AAAT

CACC

AC

OXA-

1MultiT

SO-O

for

GGCA

CCAG

ATTC

AAC

TTTC

AAG

564

MultiT

SO-O

rev

GAC

CCCA

AGTT

TCCT

GTA

AGTG

Multip

lexIICT

X-M

grou

p1grou

p2andgrou

p9

CTX-

Mgrou

p1

MultiC

TXMGp1

for

TTAG

GAART

GTG

CCGCT

GYA

688

MultiC

TXMGp1-2

rev

CGAT

ATCG

TTGGTG

GTR

CCAT

CTX-

Mgrou

p2

MultiC

TXMGp2

for

CGTT

AAC

GGCA

CGAT

GAC

404

MultiC

TXMGp1-2

rev

CGAT

ATCG

TTGGTG

GTR

CCAT

CTX-

Mgrou

p9

MultiC

TXMGp9

for

TCAAG

CCTG

CCGAT

CTGGT

561

MultiC

TXMGp9

rev

TGAT

TCTC

GCC

GCT

GAAG

CTX-

Mgrou

p825

CTX-

Mgrou

p825

CTX-

Mg825for

AAC

RCRC

AGAC

GCT

CTAC

326

CTX-

Mg825rev

TCGAG

CCGGAASG

TGTY

AT

Multip

lexIIIA

CCFOX

MOX

DHAC

ITand

EBC

ACC-

1and

ACC-

2MultiC

aseA

CCfor

CACC

TCCA

GCG

ACTT

GTT

AC346

MultiC

aseA

CCrev

GTT

AGCC

AGCA

TCAC

GAT

CC

FOX-

1toFO

X-5

MultiC

aseFOX

for

CTAC

AGTG

CGGGTG

GTT

T162

MultiC

aseFOX

rev

CTAT

TTGCG

GCC

AGGTG

AMOX-

1MOX-

2CM

Y-1CM

Y-8to

CMY-11

andCM

Y-19

MultiC

aseM

OX

for

GCA

ACAAC

GAC

AAT

CCAT

CCT

895

MultiC

aseM

OX

rev

GGGAT

AGGCG

TAAC

TCTC

CCAA

DHA-

1and

DHA-

2MultiC

aseD

HA

for

TGAT

GGCA

CAGCA

GGAT

ATTC

997

MultiC

aseD

HA

rev

GCT

TTGAC

TCTT

TCGGTA

TTCG

LAT-1toLA

T-3BIL-1CM

Y-2to

CMY-7

CMY-12

toCM

Y-18and

CMY-21

toCM

Y-23

MultiC

aseC

ITfor

CGAAG

AGGCA

ATGAC

CAGAC

538

MultiC

aseC

ITrev

ACGGAC

AGGGTT

AGGAT

AGY

ACT-1and

MIR-1

MultiC

aseEBC

for

CGGTA

AAG

CCGAT

GTT

GCG

683


Table2Antim

icrobialsusceptib

ilityprofi

leso

fES120573

L-andAmpC

-produ

cing

Ecoliiso

latesfrom

fecalsam

ples

andassociated

resistancep

atterns

Isolates

number

119864-te

stMIC

(mgL)

Resistanceg

enes

PIP

TZP

CTX

CAZ

CAZCA

LFE

PAT

MFO

XCT

TCT

TCX

TIM

IGM

AK

CICO

LTG

CFO

SEC

1gt256lt1gt256

32322

48

0032

025

lt05lt05

006

05

21lt0016

025

0016

TEM-1+C

TX-M

-15+

OXA-

1EC

2gt256lt1gt256

161605

416

0065

025

lt05lt05

012

153

025lt0016

025lt0016

TEM-1+C

TX-M

-15+

+SHV-1

EC3

gt256

64gt256gt256

gt324

12gt256

0125

025

lt05lt05

025

643

4lt0016

0125lt0016

TEM-1+C

TX-M

-15+

OXA-

1EC

4gt256

8gt256

32320064

816

0032

006

lt05lt05

006

124

4lt0016

0125lt0016

TEM-1+C

TX-M

-15+

OXA-

1EC

5gt256

32gt256

161605

412

0125

0125lt05lt05

003

83

6lt0016

025lt0016

TEM-1+C

TX-M

-15+

SHV-1

EC6

gt256

8gt256gt256

gt324

192gt256

025

025

lt05lt05

006

192

48gt32lt0016

025lt0016

TEM-1+C

TX-M

-15+

OXA-

1EC

7gt256

8gt256gt256

gt324

128

192

025

025

lt05lt05

012

128

16gt32lt0016

025lt0016

TEM-1+C

TX-M

-15+

OXA-

1EC

8gt256lt1gt256gt256

gt324

96128

025

025

lt05lt05

006

9605

025lt0016

025

0125

TEM-1+C

TX-M

-14+

OXA-

1EC

9gt256

16gt256

44006

43

30032

0032lt05lt05

006

82

025lt0016

038lt0016

TEM-1+C

TX-M

-15

EC10

gt256

4gt256

660125

46

0032

0032lt05lt05

0125

161

05lt0016

025lt0016

TEM-1+C

TX-M

-15+

SHV-1

EC11

gt256lt1gt256

48gt321

824

0032

0032lt05lt05

025

232

1lt0016

025lt0016

TEM-1+C

TX-M

-14+

OXA-

1EC


48gt256

6432

3232

025gt256

192

075lt0016

025

0016

TEM-1+C

TX-M

-15+

CMY-2+

OXA-

1EC

13gt256


64gt256

6448

gt32gt32

025

128

4gt32lt0016

038

0016

TEM-1+C

TX-M

-15+

CMY-2

PIP

piperacillin

TZP


CAZ

cefta

zidime

CAZCA

Lcefta

zidimecefta

zidime+cla

vulanicacidC

TXc

efotaxim

eFE

Pcefepime

ATMa

ztreon

amF

OX

cefoxitin

CTT

cefotetan

CTTCX


xacillin

IMIim

ipenem

GMgentamicinA

Kam

ikacinC

Iciprofl

oxacinC

OLcolistin

TGC

tigecyclin

eFO

Sfosfo

mycin



4 Discussion











Competing Interests


Acknowledgments


References




































Volume 2014

Zoology






















Advances in

Virolog y



Volume 2014




Enzyme Research



Microbiology


Table1Prim

ersu

sedfora

mplificatio

nof

thetested120573-la

ctam

aseg

enes

(Dallenn

eetal2010

[8])

PCRtype

Target

Prim

erSequ

ence

ofprim

ers(51015840-31015840

)Amplified

prod

ucts

(bp)

Multip

lexIT

EMSHV

and

OXA-

1-like

TEM

MultiT

SO-T

for

CATT

TCCG

TGTC

GCC

CTTA

TTC

800

MultiT

SO-T

rev

CGTT

CATC

CATA

GTT

GCC

TGAC

SHV

MultiT

SO-S

for

AGCC

GCT

TGAG

CAAAT

TAAAC

713

MultiT

SO-S

rev

ATCC

CGCA

GAT

AAAT

CACC

AC

OXA-

1MultiT

SO-O

for

GGCA

CCAG

ATTC

AAC

TTTC

AAG

564

MultiT

SO-O

rev

GAC

CCCA

AGTT

TCCT

GTA

AGTG

Multip

lexIICT

X-M

grou

p1grou

p2andgrou

p9

CTX-

Mgrou

p1

MultiC

TXMGp1

for

TTAG

GAART

GTG

CCGCT

GYA

688

MultiC

TXMGp1-2

rev

CGAT

ATCG

TTGGTG

GTR

CCAT

CTX-

Mgrou

p2

MultiC

TXMGp2

for

CGTT

AAC

GGCA

CGAT

GAC

404

MultiC

TXMGp1-2

rev

CGAT

ATCG

TTGGTG

GTR

CCAT

CTX-

Mgrou

p9

MultiC

TXMGp9

for

TCAAG

CCTG

CCGAT

CTGGT

561

MultiC

TXMGp9

rev

TGAT

TCTC

GCC

GCT

GAAG

CTX-

Mgrou

p825

CTX-

Mgrou

p825

CTX-

Mg825for

AAC

RCRC

AGAC

GCT

CTAC

326

CTX-

Mg825rev

TCGAG

CCGGAASG

TGTY

AT

Multip

lexIIIA

CCFOX

MOX

DHAC

ITand

EBC

ACC-

1and

ACC-

2MultiC

aseA

CCfor

CACC

TCCA

GCG

ACTT

GTT

AC346

MultiC

aseA

CCrev

GTT

AGCC

AGCA

TCAC

GAT

CC

FOX-

1toFO

X-5

MultiC

aseFOX

for

CTAC

AGTG

CGGGTG

GTT

T162

MultiC

aseFOX

rev

CTAT

TTGCG

GCC

AGGTG

AMOX-

1MOX-

2CM

Y-1CM

Y-8to

CMY-11

andCM

Y-19

MultiC

aseM

OX

for

GCA

ACAAC

GAC

AAT

CCAT

CCT

895

MultiC

aseM

OX

rev

GGGAT

AGGCG

TAAC

TCTC

CCAA

DHA-

1and

DHA-

2MultiC

aseD

HA

for

TGAT

GGCA

CAGCA

GGAT

ATTC

997

MultiC

aseD

HA

rev

GCT

TTGAC

TCTT

TCGGTA

TTCG

LAT-1toLA

T-3BIL-1CM

Y-2to

CMY-7

CMY-12

toCM

Y-18and

CMY-21

toCM

Y-23

MultiC

aseC

ITfor

CGAAG

AGGCA

ATGAC

CAGAC

538

MultiC

aseC

ITrev

ACGGAC

AGGGTT

AGGAT

AGY

ACT-1and

MIR-1

MultiC

aseEBC

for

CGGTA

AAG

CCGAT

GTT

GCG

683


Table2Antim

icrobialsusceptib

ilityprofi

leso

fES120573

L-andAmpC

-produ

cing

Ecoliiso

latesfrom

fecalsam

ples

andassociated

resistancep

atterns

Isolates

number

119864-te

stMIC

(mgL)

Resistanceg

enes

PIP

TZP

CTX

CAZ

CAZCA

LFE

PAT

MFO

XCT

TCT

TCX

TIM

IGM

AK

CICO

LTG

CFO

SEC

1gt256lt1gt256

32322

48

0032

025

lt05lt05

006

05

21lt0016

025

0016

TEM-1+C

TX-M

-15+

OXA-

1EC

2gt256lt1gt256

161605

416

0065

025

lt05lt05

012

153

025lt0016

025lt0016

TEM-1+C

TX-M

-15+

+SHV-1

EC3

gt256

64gt256gt256

gt324

12gt256

0125

025

lt05lt05

025

643

4lt0016

0125lt0016

TEM-1+C

TX-M

-15+

OXA-

1EC

4gt256

8gt256

32320064

816

0032

006

lt05lt05

006

124

4lt0016

0125lt0016

TEM-1+C

TX-M

-15+

OXA-

1EC

5gt256

32gt256

161605

412

0125

0125lt05lt05

003

83

6lt0016

025lt0016

TEM-1+C

TX-M

-15+

SHV-1

EC6

gt256

8gt256gt256

gt324

192gt256

025

025

lt05lt05

006

192

48gt32lt0016

025lt0016

TEM-1+C

TX-M

-15+

OXA-

1EC

7gt256

8gt256gt256

gt324

128

192

025

025

lt05lt05

012

128

16gt32lt0016

025lt0016

TEM-1+C

TX-M

-15+

OXA-

1EC

8gt256lt1gt256gt256

gt324

96128

025

025

lt05lt05

006

9605

025lt0016

025

0125

TEM-1+C

TX-M

-14+

OXA-

1EC

9gt256

16gt256

44006

43

30032

0032lt05lt05

006

82

025lt0016

038lt0016

TEM-1+C

TX-M

-15

EC10

gt256

4gt256

660125

46

0032

0032lt05lt05

0125

161

05lt0016

025lt0016

TEM-1+C

TX-M

-15+

SHV-1

EC11

gt256lt1gt256

48gt321

824

0032

0032lt05lt05

025

232

1lt0016

025lt0016

TEM-1+C

TX-M

-14+

OXA-

1EC


48gt256

6432

3232

025gt256

192

075lt0016

025

0016

TEM-1+C

TX-M

-15+

CMY-2+

OXA-

1EC

13gt256


64gt256

6448

gt32gt32

025

128

4gt32lt0016

038

0016

TEM-1+C

TX-M

-15+

CMY-2

PIP

piperacillin

TZP


CAZ

cefta

zidime

CAZCA

Lcefta

zidimecefta

zidime+cla

vulanicacidC

TXc

efotaxim

eFE

Pcefepime

ATMa

ztreon

amF

OX

cefoxitin

CTT

cefotetan

CTTCX


xacillin

IMIim

ipenem

GMgentamicinA

Kam

ikacinC

Iciprofl

oxacinC

OLcolistin

TGC

tigecyclin

eFO

Sfosfo

mycin



4 Discussion











Competing Interests


Acknowledgments


References




































Volume 2014

Zoology






















Advances in

Virolog y



Volume 2014




Enzyme Research



Microbiology


Table2Antim

icrobialsusceptib

ilityprofi

leso

fES120573

L-andAmpC

-produ

cing

Ecoliiso

latesfrom

fecalsam

ples

andassociated

resistancep

atterns

Isolates

number

119864-te

stMIC

(mgL)

Resistanceg

enes

PIP

TZP

CTX

CAZ

CAZCA

LFE

PAT

MFO

XCT

TCT

TCX

TIM

IGM

AK

CICO

LTG

CFO

SEC

1gt256lt1gt256

32322

48

0032

025

lt05lt05

006

05

21lt0016

025

0016

TEM-1+C

TX-M

-15+

OXA-

1EC

2gt256lt1gt256

161605

416

0065

025

lt05lt05

012

153

025lt0016

025lt0016

TEM-1+C

TX-M

-15+

+SHV-1

EC3

gt256

64gt256gt256

gt324

12gt256

0125

025

lt05lt05

025

643

4lt0016

0125lt0016

TEM-1+C

TX-M

-15+

OXA-

1EC

4gt256

8gt256

32320064

816

0032

006

lt05lt05

006

124

4lt0016

0125lt0016

TEM-1+C

TX-M

-15+

OXA-

1EC

5gt256

32gt256

161605

412

0125

0125lt05lt05

003

83

6lt0016

025lt0016

TEM-1+C

TX-M

-15+

SHV-1

EC6

gt256

8gt256gt256

gt324

192gt256

025

025

lt05lt05

006

192

48gt32lt0016

025lt0016

TEM-1+C

TX-M

-15+

OXA-

1EC

7gt256

8gt256gt256

gt324

128

192

025

025

lt05lt05

012

128

16gt32lt0016

025lt0016

TEM-1+C

TX-M

-15+

OXA-

1EC

8gt256lt1gt256gt256

gt324

96128

025

025

lt05lt05

006

9605

025lt0016

025

0125

TEM-1+C

TX-M

-14+

OXA-

1EC

9gt256

16gt256

44006

43

30032

0032lt05lt05

006

82

025lt0016

038lt0016

TEM-1+C

TX-M

-15

EC10

gt256

4gt256

660125

46

0032

0032lt05lt05

0125

161

05lt0016

025lt0016

TEM-1+C

TX-M

-15+

SHV-1

EC11

gt256lt1gt256

48gt321

824

0032

0032lt05lt05

025

232

1lt0016

025lt0016

TEM-1+C

TX-M

-14+

OXA-

1EC


48gt256

6432

3232

025gt256

192

075lt0016

025

0016

TEM-1+C

TX-M

-15+

CMY-2+

OXA-

1EC

13gt256


64gt256

6448

gt32gt32

025

128

4gt32lt0016

038

0016

TEM-1+C

TX-M

-15+

CMY-2

PIP

piperacillin

TZP


CAZ

cefta

zidime

CAZCA

Lcefta

zidimecefta

zidime+cla

vulanicacidC

TXc

efotaxim

eFE

Pcefepime

ATMa

ztreon

amF

OX

cefoxitin

CTT

cefotetan

CTTCX


xacillin

IMIim

ipenem

GMgentamicinA

Kam

ikacinC

Iciprofl

oxacinC

OLcolistin

TGC

tigecyclin

eFO

Sfosfo

mycin



4 Discussion











Competing Interests


Acknowledgments


References




































Volume 2014

Zoology






















Advances in

Virolog y



Volume 2014




Enzyme Research



Microbiology



4 Discussion











Competing Interests


Acknowledgments


References




































Volume 2014

Zoology






















Advances in

Virolog y



Volume 2014




Enzyme Research



Microbiology





Competing Interests


Acknowledgments


References




































Volume 2014

Zoology






















Advances in

Virolog y



Volume 2014




Enzyme Research



Microbiology

















Volume 2014

Zoology






















Advances in

Virolog y



Volume 2014




Enzyme Research



Microbiology

research article fecal colonization with extended-spectrum...

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