Open access peer-reviewed chapter - ONLINE FIRST

Beta-Lactamase-Producing Genes and Integrons in Escherichia coli from Diarrheal Children in Ouagadougou, Burkina Faso

Written By

René Dembélé, Wendpoulomdé A.D. Kaboré, Issiaka Soulama, Oumar Traoré, Nafissatou Ouédraogo, Ali Konaté, Nathalie K. Guessennd, David Coulibaly N’Golo, Antoine Sanou, Samuel Serme, Soumanaba Zongo, Emmanuel Sampo, Alfred S. Traoré, Amy Gassama-Sow and Nicolas Barro

Submitted: December 2nd, 2021 Reviewed: February 10th, 2022 Published: March 30th, 2022

DOI: 10.5772/intechopen.103169

IntechOpen
Benign Anorectal Disorders - An Update Edited by Alberto Vannelli

From the Edited Volume

Benign Anorectal Disorders - An Update [Working Title]

Dr. Alberto Vannelli and Dr. Daniela Cornelia Lazar

Chapter metrics overview

25 Chapter Downloads

View Full Metrics

Abstract

This study aimed to determine the resistance of diarrheagenic Escherichia coli (DEC) strains to β-lactams antibiotics and to perform the molecular characterization of extended-spectrum β-lactamases (ESBLs) and integrons genes. It was carried out from August 2013 to October 2015 and involved 31 DEC strains isolated from diarrheal stools samples collected from children less than 5 years. The identification and characterization of DEC strains were done through the standard biochemical tests that were confirmed using API 20E and polymerase chain reaction (PCR). The antibiogram was realized by the disk diffusion method, then an amplification of the β-lactamase resistance genes and integrons by PCR was done. Out of the 419 E. coli, 31 isolates (7.4%) harbored the DEC virulence genes. From these DEC, 21 (67.7%) were ESBL-producing E. coli. Susceptibility to ESBL-producing E. coli showed that the majority of isolates were highly resistant to amoxicillin (77.4%), amoxicillin-clavulanic acid (77.4%), and piperacillin (64.5%). The following antibiotic resistance genes and integron were identified: blaTEM (6.5%), blaSHV (19.4%), blaOXA (38.7%), blaCTX-M (9.7%), Int1 (58.1%), and Int3 (19.4%). No class 2 integron (Int2) was characterized. Because of the high prevalence of multidrug-resistant ESBL organisms found, there is a need of stringent pediatric infection control measures.

Keywords

  • diarrheagenic
  • E. coli
  • extended-spectrum β-lactamases
  • integron
  • Burkina Faso

1. Introduction

Antimicrobial resistance (AMR) is one of the most serious global public health threats in this century, which is especially urgent regarding antibiotic resistance in bacteria [1], particularly in Enterobacterales[2]. This phenomenon has arisen globally in both nosocomial and community settings as a consequence of widespread antibiotics’ consumption [3]. Enterobacteralesare a large order of different types of bacteria including Escherichia colithat commonly cause infections both in healthcare settings and in communities [4]. To survive the effects of antibiotics, some Enterobacteralescan produce enzymes called extended-spectrum β-lactamases (ESBLs) that break down and destroy some commonly used antibiotics, including penicillins and cephalosporins, and make these drugs ineffective for treating infections [4]. Over the last decade, many studies have reported the presence of extended-spectrum β-lactamases (ESBL)-mediated resistance in Gram-negative bacteria causing infections in patients [5, 6, 7, 8, 9]. Infections that can be caused by ESBL-producing bacteria include urinary tract infection (UTI), diarrhea, skin infections, and pneumonia [10]. Possible medications used to treat ESBL infection include carbapenems, which are useful against infections caused by E. colior Klebsiella pneumoniaebacteria, fosfomycin, β-lactamase inhibitors, non-β-lactam antibiotics, and colistin when other medications have failed to stop the ESBL infection [10]. Unfortunately, the excessive use of antibiotics, in particular β-lactams, leads to the selection of ESBL-producing strains [11]. Because of the emergence and distribution of multidrug-resistant (MDR) E. coliis complicating the treatment of various serious infections [12, 13], the World Health Organization (WHO) has long recognized the need for an improved and coordinated global effort to contain AMR [1]. The burden of AMR, including MDR, varies between the regions; however, low- and middle-income countries share a disproportionate burden due to multitude of factors embedded in the characteristics of the health system, policy, and the practice [14].

In Burkina Faso, there is an emergence of β-lactam-resistant enterobacteria, both in rural and urban areas [9, 15, 16, 17]. Otherwise, carbapenemase-encoding genes are widespread in many parts of the world [18]. According to a previous study, carbapenemase-producing Enterobacterales(CPE) remain one of the most urgent healthcare threats [2]. To this day, the ESBLs and integrons’ genes have been poorly characterized in Burkina Faso, particularly in enteric bacteria in children less than 5 years of age. However, it is imperative that bacterial isolates from underdeveloped regions undergo extensive MDR characterization to inform national strategies designed to halt the continuing spread of these dangerous pathogens [19]. Therefore, the aim of this study was to determine the resistance of diarrheagenic E. colistrains to β-lactams antibiotics and perform the molecular characterization of extended-spectrum β-lactamases (ESBL) and integrons genes among clinical DEC isolated from stools collected in children less than 5 years of age.

Advertisement

2. Methodology

2.1 Study design, area, and sample population

It is a cross-sectional study conducted in two hospitals of Ouagadougou, Burkina Faso (Paul VI and Schiphra), during August 2013 to October 2015 (Figure 1). The Paul VI hospital is located in peripheral area and the Schiphra’s hospital in the city center at the dam edge of Ouagadougou. Many patients from Ouagadougou and its surroundings attend these two healthcare centers because of the good level of healthcare. The study population comprised children below 5 years attending the hospital for treatment.

Figure 1.

Sampling sites in Ouagadougou.

The specimens were collected adhering to a standard protocol from pediatric patients below 5 years of age with acute diarrhea and who were hospitalized or visited the health centers as outpatient. Thus, children who attended the hospitals for treatment and provided assent (from parents) or consent for the study were included in the study. Any child over the age of 5 years was excluded from the study.

2.2 Sample collection and transport

Three hundred and fifteen stool samples were collected in sterile containers and transported to the laboratory of molecular biology, epidemiology, and surveillance of bacteria and viruses transmitted by food, center for research in biological, food, and nutritional sciences at the Joseph KI-ZERBO University of Ouagadougou within 24 h in a cool box at +4°C for immediate analysis.

2.3 Bacterial isolates

Isolation of E. coliwas carried out onto eosin methylene blue agar (Liofilchem, Italy), and the plates were incubated at +37°C for 18–24 h. After this stage, the suspected E. colicolonies were selected and streaked onto Mueller-Hinton agar plate (Liofilchem, Italy). Confirmation was carried out by a biochemical microbiology method based on negative urease (Bio-Rad, France), negative citrate (Liofilchem, Italy), positive indole (Bio-Rad, France), positive lactose (Liofilchem, Italy), and positive orthonitrophenyl-β-D-galactopyranoside (ONPG) (bioMerieux, France). E. colistrains isolated were confirmed by API 20E (bioMérieux, France).

The five main pathogroups of E. coli(Enteroaggregative E. coli:EAEC, Enteropathogenic E. coli: EPEC, Enteroinvasive E. coli: EIEC, Enterohemorrhagic E. coli: EHEC, and Enterotoxigenic E. coli: ETEC) were characterized by the 16-plex polymerase chain reaction (PCR) as described by Antikainen et al. [20].

2.4 Antimicrobial susceptibility testing

All identified isolates of E. coliwere treated for susceptibility testing against amoxicillin (25 μg), amoxicillin-clavulanic acid (20/10 μg), ceftriaxone (30 μg), cefotaxime (30 μg), cefepime (30 μg), cefixime (10 μg), piperacillin (75 μg), piperacillin-tazobactam (100 + 10 μg), imipenem (10 μg), and aztreonam (30 μg) (Bio-Rad, France) following disk diffusion method on Mueller-Hinton Agar (Liofilchem, Italy). Results were interpreted based on the European Committee of Antimicrobial Susceptibility Testing (EUCAST) guidelines [21]. These isolates, which were not susceptible (either resistant or intermediate) to three or more antibiotics classes, were considered as MDR [22].

2.5 Screening and confirmation of ESBL and integrons producers

A double synergy test was used for ESBL-producing strains testing. This consisted of placing disks (2–3 cm diameter) of ceftriaxone and cefotaxime around an amoxicillin-clavulanic acid disk on the bacterial plate.

For molecular characterization, DNA extraction was performed using heating method [23]. A loopful of bacterial growth from Mueller-Hinton agar (Liofilchem, Italy) plate was suspended in 1 ml of sterilized water. The mixture was boiled for 10 min at +100°C and centrifuged for 10 min at 12000 rpm at +4°C. Supernatant was then collected and used for the PCR reactions as DNA matrices. Multiplex PCR assays were performed for detecting EBLS-encoding genes (blaTEM, 𝑏𝑙𝑎SHV, 𝑏𝑙𝑎OXA, and blaCTX-M) and the presence of the class 1, class 2, and class 3 integrons from the β-lactams-resistant DEC strains. Primers (GeneCust, France) used for these amplifications are described in Table 1.

Genetic resistance supportsGenesPrimers sequence (5′ to 3′)Product size (bp)
β-Lactams resistance geneblaTEMF: ATG AGT ATT CAA CAT TTC CG1080
R: CCA ATG CTT ATT CAG TGA GG
blaSHVF: TTA TCT CCC TGT TAG CCA CC768
R: GAT TTG CTG ATT TCG CTC GG
blaOXAF: ATG AAA AAC ACA ATA CAT ATC813
R: AAT TTA GTG TGT TTA GAA TGG
blaCTX-MF: -ATG TGC AGY ACC AGT AAR GT544
R: -TGG GTR AAR TAR GTS ACC AGA
IntegronsInt1F: ATT TCT GTC CTG GCT GGC GA600
R: ACA TGT GAT GGC GAC GCA CGA
Int2F: CAC GGA TAT GCG ACA AAA AGG T806
R: GTA GCA AAC GAC TGA CGA AAT G
Int3F: GCC CCG GCA GCG ACT TTC AG600
R: ACG GCT CTG CCA AAC CTG ACT

Table 1.

List of all primers used for antibiotic ESBL genes and integrons detection.

Thermocycling conditions were as follows: 5 min at +94°C, followed by 35 amplification cycles of +94°C for 30 s, 59 ± 4°C for 60 s, and +72°C for 60 s with a final extension of +72°C for 10 min on a thermal cycler (Gene Amp 9700, Applied Biosystems). PCR products were revealed on 1.5% stained Redsaf agarose gel (Prolabo, France), after electrophoresis under UV light (Gel Logic 200).

The PCR assays were carried out in a 25-ml reaction mixture, which consisted of 2.5 μl of the supernatant added to 22.5 μl of reaction mixture. This mixture contained 5 U of Taq DNA polymerase (Accu Power, South Korea), deoxyribonucleic triphosphate (10 mM), buffer GC (10X), MgCl2 (25 mM), and PCR primers (10 μM). Thermocycling conditions were as follows: 5 min at +94°C, followed by 35 amplification cycles at +94°C for 30 s, + 59 ± 4°C for 60 s, and +72°C for 60 s with a final extension of +72°C for 10 min on a thermal cycler (AB Applied Biosystems). Following PCR, the reaction products were separated using electrophoresis in 1.5% agarose gel (weight/volume), stained with Redsaf solution (Prolabo, France), and visualized under UV light (Gel Logic 200) [23].

2.6 Statistical analysis

The Fisher’s exact test with two-tailed pof Open Epi version 7.1.2.0 was used to determine the statistical significance of the results. A pvalue of <0.05 was considered statistically significant.

Advertisement

3. Results

3.1 Prevalence of bacterial isolates

From 315 children with diarrhea, 192 stool samples were positive to one suspected E. colidetection (60.9%). Four hundred and nineteen (419) strains of E. coliwere isolated, from which 31 DEC (7.4%) were characterized. From these DEC, 21 DEC were ESBL-producing E. coli(67.7%).

3.2 Antimicrobial susceptibility

All the DEC strains tested for the 10 β-lactams antibiotics showed important resistances to the aminopenicillins. However, few cephalosporins and carbapenems were yet active on some pathotypes (Table 2).

β-Lactams subfamiliesAntibioticsPrevalence of antibiotic susceptibility N (%)DEC resistance prevalence N (%)
ResistantSensitiveEPEC (n= 8)EHEC (n= 3)EIEC (n= 4)EAEC (n= 15)ETEC (n= 1)
PenicillinsAmoxicillin24 (77.4)7 (22.6)6 (76)2 (66.6)4 (100)11 (73.3)1 (100)
Amoxicillin-clavulanic acid24 (77.4)7 (22.6)6 (76)2 (66.6)4 (100)11 (73.3)1 (100)
Piperacillin20 (64.5)11 (35.5)5 (62.5)2 (66.6)3 (75)9 (60)1 (100)
Piperacillin-tazobactam12 (38.7)19 (61.3)3 (37.5)2 (66.6)1 (25)5 (33.3)1 (100)
CephalosporinsCeftriaxone13 (41.9)18 (58.1)2 (25)1 (33.3)2 (50)7 (46.6)1 (100)
Cefixime13 (41.9)18 (58.1)2 (25)1 (33.3)2 (50)7 (46.6)1 (100)
Cefotaxim14 (45.2)17 (54.8)2 (25)2 (66.6)2 (50)7 (46.6)1 (100)
Cefepim14 (45.2)17 (54.8)2 (25)2 (66.6)2 (50)7 (46.6)1 (100)
MonobactamAztreonam14 (45.2)17 (54.8)2 (25)2 (66.6)2 (50)7 (46.6)1 (100)
CarbapenemsImipenem5 (16.1)26 (83.9)1 (12.5)1 (33.3)0 (0)3 (20)0 (0)

Table 2.

Antimicrobials susceptibility of the studied isolates to β-lactams.

3.3 Correlation between resistance phenotype and resistance genetic supports

Nineteen (19) out of the 21 ESBLs-producing E. coli(90.5%) had ESBLs genes. The following resistance genes were characterized: 12 blaOXA (38.7%), 6 blaSHV (19.4%), 3 𝑏𝑙𝑎CTX-M (9.7%), and 2 blaTEM (6.5%). Our results showed that the genes responsible for the production of blaOXA β-lactamases 12/31 (38.7%) were more prevalent in comparison to the genes encoding blaTEM, blaSHV, and 𝑏𝑙𝑎CTX-M β-lactamases (Table 3). From the three classes of integrons (Int1, Int2, and Int3) assessed among the resistant strains carrying ESBL genes, only 18 Int1(58.1%) and 2 Int3(19.4%) were detected. The class 3 integron was detected in only EIEC. No class 2 integrons (Int2) were characterized from the resistant strains. The coexistence of the three resistance genes (blaSHV, blaOXA, and blaCTX-M) and Int1was found in one EHEC (Table 3). The blaOXA gene (Figure 2) was associated with Int1(Figure 3) in 11 cases (p = 0.001), while the blaSHV gene was associated with Int1in 5 cases (p = 0.100).

DEC strainsResistance phenotypesAntibioticsGenetic resistance supports
Resistance genesIntegrons
AMCAMXCTXATMIPMCROFEPCFMTZPPIPblaTEMblaSHVblaOXAblaCTX-MIntI1IntI2IntI3
Schiphra’s hospital (n = 9/18; 50%)
EHECESBL, CarbapenemaseRRRRR-R-RR-++++--
aEPECESBLRR--------+---+--
tEPECESBLRRRR-RRRRR--+-+--
tEPECESBLRR------RR-+--+--
EIECESBLRR-------R-+--+-+
EAECESBLRR-------R-+--+--
EAECESBLRR---------+--+--
EAECESBL, CarbapenemaseRRRRRRRRRR---++--
EAECESBLRRRR-RRRRR--+-+--
Paul VI hospital (n = 10/13; 76.9%)
aEPECESBL, PHNRR-------R--+-+--
aEPECESBL, CarbapenemaseRRRRRRRRRR--+-+--
aEPECESBL, PHNRR-------R--+-+--
EIECESBL, PHNRRRR-RRRRR-++---+
EAECESBL, PHNRR-------R--+-+--
ETECCASE, PHNRRRR-RRRRR--+-+--
EAECESBL, PHNRR-------R--+-+--
EAECESBL, PBNRRRR-RRR-S+---+--
EAECESBL, CarbapenemaseRRRRRRRRRR--+++--
EAECESBLRRRR-RRR----+-+--

Table 3.

Correlation between E. colipathotypes, antibiotics resistance, and genetic resistance supports.

EAEC = Enteroaggregative E. coli, aEPEC = Atypical Enteropathogenic E. coli, tEPEC = Typical Enteropathogenic E. coli, EIEC = Enteroinvasive E. coli, EHEC = Enterohemorrhagic E. coli, ETEC = Enterotoxigenic E. coli, PHN = High-level penicillinases, CASE = Cephalosporinases, ESBL = Extended-spectrum β-lactamases, AMC = Amoxicillin-Clavulanic Acid, AMX = Amoxicillin, CTX = Cefotaxime, ATM = Aztreonam, IPM = Imipenem, CRO = Ceftriaxone, FEP = Cefepime, CFM = Cefixime, TZP = Piperacillin-tazobactam, PIP = Piperacillin, R = Resistant, − = Absence, + = Presence.

Figure 2.

blaOXA gene on agarose gel electrophoresis (1.5%). Lane M: molecular size marker (100 bp), 1:blaOXA1: positive control (813 pb), lanes: 2–8 are positive forblaOXA gene, lane T: negative control.

Figure 3.

Int1 gene on agarose gel electrophoresis (1.5%). Lane A: molecular size marker (100 bp), B:Int1: positive control (600 pb), lanes: C-O are positive forInt1gene, lane T: negative control.

Advertisement

4. Discussion

The emergence and spread of multidrug-resistant (MDR) bacteria are major public health threats worldwide. Particularly, DEC that produce ESBL are of great concern, because their resistance to penicillins and narrow extended-spectrum cephalosporins reduces considerably the treatment options. The prevalence of ESBL in Enterobacteriaceaehas been detected at local levels in various African countries; moreover, a study was conducted in 2014 on the prevalence of ESBL and what type of genes are involved in its occurrence [24]. The frequency of ESBL-producing E. coliwas 67.7% in our study. Similar prevalence was reported in Egypt (69.6%) [25] and Palestine (66.7%) [26]. Nevertheless, our prevalence was higher than those in Burkina Faso (58%) [6], Iran (40.8%) [27], Saudi Arabia (30.6%) [28], Japan (20.4%) [29], Colombia (11.7%) [30], and Nepal (22.7%) [31]. Otherwise, our result is lower than the ESBL production in clinical isolates of E. colireported somewhere else in Iran [32]. The prevalence of ESBL resistance in E. coliisolates in European countries is reported to be around 3.9% with variations between countries [33]. Overall, these percentages are lower than those found in middle-income countries like Thailand (71.25%) [34] and China (50.5%) [35]. This difference between ESBLs’ prevalence might be due to patient’s age, the type of samples, and the country health facilities in the management of diarrheal infections regarding antibiotics use. Indeed, in developing countries, most patients received antibiotics treatment without prescription [36, 37]; such common practices in nearly all developing countries cause a selective pressure on E. coli, whereas in more developed countries effective strategies for the control of antimicrobial are present, which effectively prevents the emergence of ESBLs [36].

It has been reported that bacteria such as E. coliand K. pneumoniaeare major ESBL producers resulting in serious threat to the treatment regimen [38]. Indeed, ESBL enzymes are becoming increasingly expressed by many strains of pathogenic bacteria presenting diagnostic challenges to the clinical microbiology laboratories [39, 40]. Until recently, antimicrobial therapy has played an important role in the treatment of human bacterial infections. However, the drug resistance has emerged in the treatment of bacterial infections due to ESBL enzymes [39]. Indeed, these enzymes can degrade all β-lactam antibiotics leading to multidrug-resistant bacteria. Therefore, reporting of ESBL-producing isolates from clinical samples is critical for the clinicians. It constitutes the guidelines to select appropriate antibiotics for the treatment, including to take proper precaution to prevent the spread of these resistant organisms to other patients [31].

The present study shows 19 ESBLs genes (90.5%) out of the 21 ESBLs-producing E. coli. Analysis of the ESBL-encoding genes indicated that the majority of the ESBL-positive isolates harbored blaOXA (38.7%), followed by blaSHV (19.4%), blaCTX-M (9.7%), and 𝑏𝑙𝑎TEM (6.5%). The emergence of β-lactam resistance in Enterobacteriaceaeis related primarily to the production of enzymes such as TEM and SHV variant, which were the most common ESBLs during the past decade. However, OXA and CTX-M β-lactamases have emerged as prevalent ESBL worldwide type compared with the TEM and SHV genotypes [41].

In the present study, OXA-type ESBL-producing DEC strains (38.7%) were the most frequently detected ESBL gene. This prevalence is lower than that reported in our previous study in rural area of Burkina Faso: 100% [9], also lower comparatively to 52% reported in Pakistan [42]. However, a recent study in young children reported 3% of commensal E. colibearing the blaOXA gene in Bangladesh [41]. Thus, it appears that the emergence of ESBLs-producing bacteria among gut bacteria of young children can transfer resistance and related genes horizontally across pathogenic E. coli, and commensal E. colileading to a public health concern. Most of the OXA-type ESBL-producing E. coliisolates (29%) in our study were detected from the Paul VI hospital (p = 0.002). This hospital is located in peripheral area of Ouagadougou, and most of the people living in the slums with poor sanitation conditions attend it for healthcare sought. Moreover, the provision of confessional care has less difficult accessibility for the peripheral neighborhoods and the population with low socioeconomic level. Otherwise, people in Burkina Faso do not consult a healthcare agent in the case of diseases such as gastrointestinal infections and use self-medication instead [37]. Our results showed 19.4% of SHV-type ESBL-producing E. coliwhich is a little similar to 21% detected in Pakistan [42]. By cons, this prevalence is higher than 0% [9] and 5.9% [17], previously reported in Burkina Faso but lower than 45% reported in Iran [27]. The blaCTX-M gene (Figure 4) has been detected in three E. coliisolates, while its prevalence was 25% in our earlier report [9] and 40.1% by a study conducted in Enterobacteriaceaefrom Burkinabe patients [17]. Moreover, few studies from other parts of world have shown different prevalence of blaCTX-M gene among isolates, including 98.8% (China), 84.7% (Chile), 13.6% (Tanzania), 76% (Pakistan), 97.8% (Chad), and 81.6% (Egypt) [25, 42, 43, 44, 45, 46]. Indeed, CTX-M β-lactamases are recognized as the most widespread extended-spectrum β-lactamases (ESBLs) among clinical isolates of Enterobacteriaceae[47]. Besides, an earlier report from Nigeria has shown the predominance of CTX-M15 in wild birds and cattle in Nigeria [48] suggesting that this gene could be transferred to humans by animals. Finally, our study revealed 6.5% of TEM-type ESBL-producing E. coli, while no blaTEM gene has been detected in our previous study [9]. However, this value is lower than 26.2 and 28% reported in Burkina Faso and Pakistan, respectively [17, 42]. The resistance to amoxicillin/amoxicillin-clavulanic acid observed in the two E. colistrains (6.5%) may be mainly mediated by the production of these plasmid-encoded TEM enzymes.

Figure 4.

blaCTX-M gene on agarose gel electrophoresis (1.5%). Lane M: molecular size marker (100 bp), A:blaCTX-Mpositive control (544 pb), lanes: B, D, and E are positive forblaCTX-Mgene (544 pb), lane T: negative control.

Among the three class of integron, class 1 integron (58.1%) was majority characterized from the resistant strains in accordance with 56% reported in Bangladesh [41]. This result confirms those of previous studies showing that class 1 integron was predominantly represented in Enterobacteriaceae[49, 50]. However, a previous report in Burkina Faso has shown a lower prevalence (44.4%) of Int1[51]. On the other hand, studies reported a high prevalence of Int1(80%) in E. coliisolated from dairy products consumed in Burkina Faso [52] and in human, animal, and food in Spain [53]. This could increase the risk of emergence and spread of MDR E. coli, since humans are always in contact with these different ecosystems, especially when there is a lack of food hygiene and sanitation. Moreover, class 1 integrons can facilitate the spread of antibiotic-resistant genes meaning that it could have public health consequences [54].

The class 3 integron was detected in only EIEC. No class 2 integron (Int2) was characterized from the resistant strains. By cons, 22.2% of Int2was detected in our previous study [51]. Moreover, a study also found the presence of Int2gene in Senegalese Shigellaspp. isolates [49].

Two strains of EIEC harbored both class 1 and 3 integrons. However, a previous study showed that E. coliharbored class 1 and 2 integrons simultaneously [50]. Otherwise, in the present study, one EIEC strain was resistant to aztreonam and imipenem and possesses ESBL-carbapenemase phenotype. This strain was resistant to all subfamilies (penicillins, cephalosporins, monobactam, and carbapenems) of β-lactams antibiotic tested and also showed simultaneous presence of blaSHV, blaOXA,blaCTX-M, and Int1. Indeed, strains that had this aztreonam-resistant phenotype possessed both the resistance gene [27]. Resistance to this antibiotic could be explained by genetic mutations [43]. It has been described that the coexistence of these two classes of integrons [42] and/or several genes suggests that they have integrated the same gene and give these strains a high level of resistance. However, blaTEM, blaSHV, blaOXA, blaCTX-M as well as integrons (Int1, Int2, and Int3) are involved in the antibiotic resistance of DEC, but the presence of resistant strains producing ESBL and lacking ESBL gene (blaTEM, blaSHV, blaOXA, and blaCTX-M) and integron suggests that there are other mechanisms for the dissemination of antibiotic resistance in DEC strains.

Advertisement

5. Conclusion

This study highlights the important involvement of genes and integrons into multidrug resistance strains of E. coliin two main hospitals of Ouagadougou. The most important finding was the detection of four E. colimultiresistant strains producing ESBL that were resistant to imipenem, aztreonam, and harbored class 1 integrons. Another important observation was the detection of two E. colimultiresistant strains producing ESBL but lacking a resistance gene and/or integrons. Our results have demonstrated the emergence and dissemination of multidrug-resistant E. colistrains hosting several genes responsible for the production of ESBL in clinical isolates. Ultimately, to fight effectively against the emergence of antimicrobial resistance, an integrated surveillance network should be set up, which would be of great benefit to national antimicrobial resistance control programs.

Advertisement

Acknowledgments

The authors gratefully thank “Réseau de Recherche sur les Maladies Entériques à potentiel épidémique en Afrique de l’Ouest (REMENTA)/Programme d’Appui à la Recherche en Réseau en Afrique (PARRAF)”for technical support. The authors also thank the parents and guardians of children as well as the authorities of the Paul VI and Schiphra’s hospitals for their honest cooperation.

References

  1. 1. Prestinaci F, Pezzotti P, Pantosti A. Antimicrobial resistance: A global multifaceted phenomenon.Pathogens and Global Health. 2015;109(7):309-318
  2. 2. Dembélé R, Soulama I, Kaboré WAD, Konaté A, Kagambèga A, Coulibaly DN, et al. Molecular characterization of carbapenemase-producingEnterobacteralesin children with diarrhea in rural Burkina Faso. Journal of Drug Delivery and Therapeutics. 2021;11(1):84-92
  3. 3. Ashley EA, Lubell Y, White NJ, Turner P. Antimicrobial susceptibility of bacterial isolates from community acquired infections in Sub-Saharan Africa and Asian low and middle income countries. Tropical Medicine & International Health. 2011;16:1167-1169
  4. 4. Centers for Disease Control and Prevention, National Center for Emerging and Zoonotic Infectious Diseases (NCEZID), Division of Healthcare Quality Promotion (DHQP). Healthcare-Associated Infections (HAI),Diseases and Organisms. 2019
  5. 5. Raut S, Gokhale S, Adhikari B. Prevalence of extended spectrum beta-lactamases amongE. coliandKlebsiellaspp isolates in Manipal, Teaching Hospital, Pokhara, Nepal. Journal of Microbiology and Infectious Diseases. 2015;5:69-75
  6. 6. Ouédraogo AS, Sanou M, Kissou A, Sanou S, Solaré H, Kaboré F, et al. High prevalence of extended-spectrum β-lactamase producingEnterobacteriaceaeamong clinical isolates in Burkina Faso. BMC Infectious Diseases. 2016;16:326
  7. 7. Nepal K, Pant ND, Neupane B, Belbase A, Baidhya R, Shrestha RK, et al. Extended spectrum beta-lactamase and metallo beta-lactamase production amongEscherichia coliandKlebsiella pneumoniaeisolated from different clinical samples in a tertiary care hospital in Kathmandu, Nepal. Annals of Clinical Microbiology and Antimicrobials. 2017;16:62
  8. 8. Rai S, Pant ND, Bhandari R, Giri A, Parajuli R, Aryal M, et al. AmpC and extended spectrum beta-lactamases production among urinary isolates from a tertiary care hospital in Lalitpur, Nepal. BMC Research Notes. 2017;10:467
  9. 9. Dembélé R, Konaté A, Traoré O, Kaboré WAD, Soulama I, Kagambèga A, et al. Extended spectrum beta-lactamase and fluoroquinolone resistance genes amongEscherichia coliandSalmonellaisolates from children with diarrhea, Burkina Faso. BMC Pediatrics. 2020a;20:459
  10. 10. Jewell T, Biggers A. ESBLs (Extended Spectrum Beta-Lactamases). 2017. Available from:https://www.healthline.com/health/esbl[Updated: 14 April]
  11. 11. Chang YT, Coombs G, Ling T, Balaji V, Rodrigues C, Mikamo H, et al. Epidemiology and trends in the antibiotic susceptibilities of Gram negative bacilli isolated from patients with intra-abdominal infections in the Asia-Pacific region, 2010-2013. International Journal of Antimicrobial Agents. 2017;49:734-739
  12. 12. Allocati N, Masulli M, Alexeyev MF, Ilio CD.Escherichia coliin Europe: An overview. International Journal of Environmental Research and Public Health. 2013;10:6235-6254
  13. 13. Mariappan S, Sekar U, Kamalanathan A. Carbapenemase-producingEnterobacteriaceae: Risk factors for infection and impact of resistance on outcomes. International Journal of Applied & Basic Medical Research. 2018;7:32-39
  14. 14. Pokharel S, Raut S, Adhikari B. Tackling antimicrobial resistance in low- and middle-income countries. BMJ Global Health. 2019;4:e002104
  15. 15. Dembélé R, Bonkoungou IJO, Konaté A, Bsadjo-Tchamba G, Bawa HI, Bako E, et al. Serotyping and antibiotic resistance of enteropathogenicEscherichia coliandE. coliO157 isolated from diarrheal children in rural area of Burkina Faso. African Journal of Microbiology Research. 2015;9:1053-1059
  16. 16. Ouédraogo A-S, Sanou S, Kissou A, Poda A, Aberkane S, Bouzinbi N, et al. Fecal carriage ofEnterobacteriaceaeproducing extended-spectrum beta-lactamases in hospitalized patients and healthy community volunteers in Burkina Faso. Microbial Drug Resistance. 2017;23:1
  17. 17. Kpoda DS, Ajayi A, Somda M, Traore O, Guessennd N, Ouattara AS, et al. Distribution of resistance genes encoding ESBLs inEnterobacteriaceaeisolated from biological samples in health centers in Ouagadougou, Burkina Faso. BMC Research Notes. 2018;11:471
  18. 18. Halat DH, Moubareck CA. The current burden of carbapenemases: Review of significant properties and dissemination among gram-negative bacteria. Antibiotica. 2020;9:186
  19. 19. Margulieux KR, Srijan A, Ruekit S, Nobthai P, Poramathikul K, Pandey P, et al. Extended-spectrum β-lactamase prevalence and virulence factor characterization of enterotoxigenicEscherichia coliresponsible for acute diarrhea in Nepal from 2001 to 2016. Antimicrobial Resistance and Infection Control. 2018;7:87
  20. 20. Antikainen J, Tarkka E, Haukka K, Siitonen A, Vaara M, Kirveskari J. New 16 plex PCR method for rapid detection of diarrheagenicEscherichia colidirectly from stool samples. European Journal of Clinical Microbiology & Infectious Diseases. 2009;28:899-908
  21. 21. European Committee on Antimicrobial Susceptibility Testing (EUCAST). Recommandation 2017. Éd. V.1.0 Mars. 2017. pp. 1-127. Available from:https://www.eucast.org/
  22. 22. Magiorakos AP, Srinivasan A, Carey RB, Carmeli Y, Falagas ME, Giske CG, et al. Multidrug-resistant, extensively drug-resistant and pandrug-resistant bacteria: An international expert proposal for interim standard definitions for acquired resistance. Clinical Microbiology and Infection. 2012;18:268-281
  23. 23. Moyo SJ, Maselle SY, Matee MI, Langeland N, Mylvaganam H. Identification of diarrheagenicEscherichia coliisolated from infants and children in Dar es Salaam, Tanzania. BMC Infectious Diseases. 2007;7(92):1-7
  24. 24. Storberg V. ESBL-producingEnterobacteriaceaein Africa—A non-systematic literature review of research published 2008-2012. Infection Ecology & Epidemiology. 2014;4(1):20342
  25. 25. Mohamed ES, Khairy RMM, Abdelrahim SS. Prevalence and molecular characteristics of ESBL and AmpC β-lactamase producingEnterobacteriaceaestrains isolated from UTIs in Egypt. Antimicrobial Resistance and Infection Control. 2020;9:198
  26. 26. Tayh G, Laham NA, Yahia HB, Sallem RB, Elottol AE, Slama KB. Extended-spectrum β-lactamases among enterobacteriaceae isolated from urinary tract infections in Gaza Strip, Palestine. BioMed Research International. 2019. Article ID: 4041801,11
  27. 27. Seyedjavadi SS, Goudarzi M, Sabzehali F. Relation betweenblaTEM,blaSHV andblaCTX-M genes and acute urinary tract infections. Journal of Acute Diseases. 2016;5:71-76
  28. 28. Hassan H, Abdalhamid B. Molecular characterization of extended spectrum beta-lactamase producingEnterobacteriaceaein a Saudi Arabian tertiary hospital. Journal of Infection in Developing Countries. 2014;8:282-288
  29. 29. Harada Y, Morinaga Y, Yamada K, Migiyama Y, Nagaoka K, Uno N, et al. Clinical and molecular epidemiology of extended spectrum β-lactamase-producingKlebsiella pneumoniaeandEscherichia coliin a Japanese tertiary hospital. Journal of Medical Microbiology and Diagnosis. 2013;2:127
  30. 30. Martinez P, Garzôn D, Mattar S. CTX-M-producingEscherichia coliandKlebsiella pneumoniaeisolated from community-acquired urinary tract infections in Valledupar, Colombia. Brazilian Journal of Infectious Diseases. 2012;16:420-425
  31. 31. Kayastha K, Dhungel B, Karki S, Adhikari B, Banjara MR, Rijal KR, et al. Extended-spectrum β-lactamase-producingEscherichia coliandKlebsiellaspecies in pediatric patients visiting International Friendship Children’s Hospital, Kathmandu, Nepal. Infectious Diseases: Research and Treatment. 2020;13:1-7
  32. 32. Najar PS, Eslami M, Memariani M, Siadat SD. High prevalence ofblaCTX-M-1 group extended spectrum β-lactamase genes inEscherichia coliisolates from Tehran. Jundishapur Journal of Microbiology. 2013;6:e6863
  33. 33. Ahmed MO, Clegg PD, Williams NJ, Baptiste KE, Bennett M. Antimicrobial resistance in equine faecalEscherichia coliisolates from North West England. Annals of Clinical Microbiology and Antimicrobials. 2010;9:12
  34. 34. Luvsansharav UO, Hirai I, Niki M, Sasaki T, Makimoto K, Komalamisra C, et al. Analysis of risk factors for a high prevalence of extended-spectrum βlactamase-producingEnterobacteriaceaein asymptomatic individuals in rural Thailand. Journal of Medical Microbiology. 2011;60:619-624
  35. 35. Li B, Sun JY, Liu QZ, Han LZ, Huang XH, Ni YX. High prevalence of CTX-M β-lactamases in faecalEscherichia colistrains from healthy humans in Fuzhou, China. Scandinavian Journal of Infectious Diseases. 2011;43:170-174
  36. 36. Ahmed SF, Ali MMM, Mohamed ZK, Moussa TA, Klena JD. Fecal carriage of extended-spectrum β-lactamases and AmpC-producingEscherichia coliin a Libyan Community. Annals of Clinical Microbiology and Antimicrobials. 2014;13:22
  37. 37. Dembélé R, Huovinen E, Yelbéogo D, Kuusi M, Sawadogo G, Haukka K, et al. Burden of acute gastrointestinal infections in Ouagadougou, Burkina Faso. Journal of Microbiology Infectious Diseases. 2016;6:45-52
  38. 38. Rimal U, Thapa S, Maharajan R. Prevalence of extended spectrum beta-lactamase producingEscherichia coliandKlebsiellaspecies from urinary specimens of children attending Friendship International Children’s Hospital. Nepal Journal of Biotechnology. 2017;5:32-38
  39. 39. Gupta V. An update on newer β-lactamases. The Indian Journal of Medical Research. 2007;126:417-427
  40. 40. Sharma AR, Bhatta DR, Shrestha J, Banjara MR. Antimicrobial susceptibility pattern ofEscherichia coliisolated from urinary tract infected patients attending Bir Hospital. Nepal Journal of Science and Technology. 2013;14:177-184
  41. 41. Monira S, Shabnam SA, Imran ASK, Sadique A, Johura FT, Rahman KZ, et al. Multi-drug resistant pathogenic bacteria in the gut of young children in Bangladesh. Gut Pathogenesis. 2017;9:19
  42. 42. Abrar S, Ul AN, Liaqat H, Hussain S, Rasheed F, Riaz S. Distribution ofblaCTX-M,blaTEM,blaSHV andblaOXA genes in extended-spectrum βlactamase-producing clinical isolates: A three-year multi-center study from Lahore, Pakistan. Antimicrobial Resistances and Infection Control. 2019;8:80
  43. 43. Quan J, Zhao D, Liu L, Chen Y, Zhou J, Jiang Y, et al. High prevalence of ESBL producingEscherichia coliandKlebsiella pneumoniaein community-onset bloodstream infections in China. The Journal of Antimicrobial Chemotherapy. 2016;72(1):273-280
  44. 44. Sonda T, Kumburu H, van Zwetselaar M, Alifrangis M, Mmbaga BT, Lund O, et al. Prevalence and risk factors for CTX-M gram-negative bacteria in hospitalized patients at a tertiary care hospital in Kilimanjaro, Tanzania. European Journal of Clinical Microbiology & Infectious Diseases. 2018;37:1-10
  45. 45. Pavez M, Troncoso C, Osses I, Salazar R, Illesca V, Reydet P, et al. High prevalence of CTX-M-1 group in ESBL-producingEnterobacteriaceaeinfection in intensive care units in southern Chile. The Brazilian Journal of Infectious Diseases. 2019;23(2):102-110
  46. 46. Ouchar Mahamat O, Tidjani A, Lounnas M, Hide M, Benavides J, Somasse C, et al. Fecal carriage of extended-spectrum βlactamase-producingEnterobacteriaceaein hospital and community settings in Chad. Antimicrobial Resistance and Infection Control. 2019;8:169
  47. 47. Tani BA-KZ, Decré D, Genel N, Boucherit-Otmani Z, Arlet G, Drissi M. Molecular and epidemiological characterization of enterobacterial multidrug-resistant strains in Tlemcen Hospital (Algeria) (2008-2010). Microbial Drug Resistance. 2013;19(3):185-190
  48. 48. Fashae K, Engelmann I, Monecke S, Braun DS, Ehricht R. Molecular characterisation of extended spectrum ß-lactamase producingEscherichia coliin wild birds and cattle, Ibadan, Nigeria. BMC Veterinary Research. 2021;17:33
  49. 49. Gassama-Sow A, Aïdara-Kane A, Barraud O, Gatet M, Denis F, Ploy MC. High prevalence of trimethoprim-resistance cassettes in class 1 and 2 integrons in SenegaleseShigellaspp isolates. Journal of Infection in Developing Countries. 2010;4:207-212
  50. 50. Sambe-Ba B, Seck A, Wane AA, Fall-Niang NK, Gassama-Sow A. Sensibilité aux antibiotiques et supports génétiques de la résistance des souches deShigella flexneriisolées à Dakar de 2001 à 2010. Bulletin de la Societe de Pathologie Exotique. 2013;106:89-94
  51. 51. Dembélé R, Kaboré WAD, Soulama I, Konaté A, Kagambèga A, Traoré O, et al. Involvement of class 1 and class 2 integrons in dissemination oftetandcatA1 resistance genes ofSalmonella entericafrom children with diarrhea in rural Burkina Faso. African Journal of Biotechnology. 2020b;19(1):1-7
  52. 52. Bagré TS, Sambe-Ba B, Bawa-Ibrahim H, Bsadjo-Tchamba G, Dembélé R, Wane AA, et al. Isolation and characterization of enteropathogenic and enterotoxinogenicEscherichia colifrom dairy products consumed in Burkina Faso. African Journal of Microbiology Research. 2017;11(13):537-545
  53. 53. Sáenz Y, Briñas L, Domínguez E, Ruiz J, Zarazaga M, Vila J, et al. Mechanisms of resistance in multiple-antibiotic-resistantEscherichia colistrains of human, animal, and food origins. Antimicrobial Agents and Chemotherapy. 2004;48:3996-4001
  54. 54. Oliveira-Pinto C, Diamantino C, Oliveira PL, Reis MP, Costa PS, Paiva MC, et al. Occurrence and characterization of class 1 integrons inEscherichia colifrom healthy individuals and those with urinary infection. Journal of Medical Microbiology. 2017;66(5):577-583

Written By

René Dembélé, Wendpoulomdé A.D. Kaboré, Issiaka Soulama, Oumar Traoré, Nafissatou Ouédraogo, Ali Konaté, Nathalie K. Guessennd, David Coulibaly N’Golo, Antoine Sanou, Samuel Serme, Soumanaba Zongo, Emmanuel Sampo, Alfred S. Traoré, Amy Gassama-Sow and Nicolas Barro

Submitted: December 2nd, 2021 Reviewed: February 10th, 2022 Published: March 30th, 2022