Acta Scientific Microbiology

Research Article Volume 1 Issue 1

Forward and Reverse Characterization of the CTX-m Genes Associated with Multi-Drug Resistant Escherichia coli Isolated from Pregnant Mothers Presenting with Asymptomatic Urinary Tract Infection in Benin City, Nigeria

Otabor Florence Enogiomwan1* and Isaiah Nnanna Ibeh2

1Health Services Department, University of Benin, Benin City, Edo state, Nigeria
2Department of Medical Laboratory Sciences, Faculty of Basic Medical Sciences, University of Benin city Edo state, Nigeria

*Corresponding Author: Otabor Florence Enogiomwan, Health Services Department, University of Benin, Benin City, Edo state, Nigeria.

Received: December 04, 2017; Published: December 22, 2017

DOI: 10.31080/ASMI.2018.01.0005

Citation: Otabor Florence Enogiomwan and Isaiah Nnanna Ibeh. “Forward and Reverse Characterization of the CTX-m Genes Associated with Multi-Drug Resistant Escherichia coli Isolated from Pregnant Mothers Presenting with Asymptomatic Urinary Tract Infection in Benin City, Nigeria”. Acta Scientific Microbiology 1.1 (2018).


Introduction: The determination and elimination of multi drug resistant Escherichia coli at the maternal levels which can culmi- nate in reduction of fetal outcomes amongst neonates.

Aim of Study: This study was aimed at determining the forward and reverse characteristics of the CTXm genes associated with multi drug resistant Escherichia coli amongst pregnant mothers presented at the anti-natal clinics in Benin city, Nigeria.

Material and Method: A total of 530 vaginal swabs were collected cross sectional from pregnant women at the different trimester of pregnancy. Samples were screened and analyzed microbiological and susceptibility profiles carried out on the isolates confirmed to be Escherichia coli using the standard Kirby-Bauer, method, both phenotypic and genetic characterization was done, using the double disc synergy test for ESBL and reverse and forward characterization of the drug resistant plasmid CTX-M.

Result and Conclusion:The result of the study revealed that (47) 8.9% from third trimester out of the 530 Samples were positive for E. coli , out of the 17 positive E. coli isolates, 14 carried CTX-M genes from the forward and reverse characterization. The detec- tion of CTX-M gene was more common in the third trimester when compared with the three trimesters. Vaginal Escherichia coli in the third trimester is challenging because they can be transferred to the neonate which can result in septicemia and pyrexia.

Keywords: Characterization; Septicemia; Susceptibility


   Escherichia coli has a place with the group of the Enterobacte- riaceae with the family Escherich which was named after its author Theodore Escherich who performed the spearheading thinks about on the fecal vegetation and portrayed the living being in 1885 [2,3] in spite of the fact that they have the quality which encodes for cephalosporinase, ampC [1] , the apparently wild kind of E. coli are defenseless to most β-lactams because of the nonappearance of a viable ampC promoter district. The predictable utilization of the β-lactam anti-infection agents birthed the development of safe strains around the world. This sort of medication protection is for the most part achieved by the procurement of β-lactamase qualities which is situated on transposable components such as plasmids. The most widely recognized β-lactamases found in E. coli have a place with class An Ambler and can be partitioned into tight and Broad range β-lactamases (e.g. TEM-1, TEM-2, and SHV-1, TEM-3, SHV-5, and CTX-M-like) (1– 3). Stretched out betalactamases pres - ent protection from the cephalosporin’s, broadly used to treat E. coli contamination.

   The World Health Organization (WHO) defines traditional medi - cine as: “the health practices, approaches, knowledge and beliefs incorporating plant, animal and mineral based medicines, spiritual therapies, manual techniques and exercises, applied singularly or in combination to treat, diagnose and prevent illnesses or maintain well-being [2] .

  Referred to CTX-Ms are apparently observed as cefotaximases that typically hydrolyze cefotaxime other than ceftazidime, not- withstanding, now and then point transformations can stretch out their objective to incorporate ceftazidime, CTX-M-15 and CTX- M-27 are gotten by substitution of Asp240Gly separately from CTX- M-3 and CTX-M-14 [7,8] . Betalactamase of the CTX-M– sort are essentially wide range β-lactamases gotten from the encoding of β-lactamases chromosomally of Kluyvera sp [4-6] . More than sev - enty sorts of CTX-M have so far been disengaged and isolated into 5 groups in view of their amino corrosive succession: CTX-M-1, CTX- M-2, CTX-M-8, CTX-M-9, and CTX-M-25.

   carcely any bacteriologic information are accessible for Cam- bodia, and no investigations of protection of Enterobacteriaceae to antimicrobial operators have been accounted for. We examined E. coli CA-UTIs in a 2-year imminent examination. Our points were 1) to build up the predominance of group procured urinary E. coli protection from an extensive variety of antimicrobial medications and 2) to describe the instruments basic E. coli protection from β-lactams.

Materials and Methods

Test gathering

  Vaginal swabs were gathered u sing sterile cotton swab with- out utilizing a speculum from the lower vaginal divider [11] . The swabs were promptly set into a vehicle medium (stuarts transport medium) and transported to the research facility.

Distinguishing proof of Isolate

   Tests were microbiologically dissected utilizing standard asep- tic condition utilizing Mac conkey agar, blood agar which was brooded vigorously at 370C for 24 hours. Secludes where related to the standard microbiological methods [9,10] .

   This was conveyed asceptically utilizing the standard Kirby buer strategy utilizing Mueller Hinton agar routinely crosswise over different expansive multi drugs barrel, which was hatched at 370C for 24 hours, zones of restraints where deciphered after Incu- bation, they were recorded as delicate, transitional vulnerable and safe as per the CLSI interpretative measures for the enterobacteria - ceae (CLSI, 2010).

Plasmid DNA Extraction

  Enhancement items were decontaminated with Montage PCR Filter Units (Millipore, Billerica, MA, USA). Sequencing responses were performed in a PTC-225 Peltier Thermal Cycler (MJ Research, Waltham, MA, USA) by utilizing an ABI PRISM BigDye Terminator Cycle Sequencing Kit with AmpliTaq DNA polymerase (Applied Bio - systems, Branchburg, NJ, USA), as indicated by the maker’s direc- tions. Every format was sequenced with the proper groundwork. Fluorescence-marked pieces were sanitized from the unincorpo - rated eliminators with an ethanol precipitation convention. The specimens were resuspended in refined water and subjected to electrophoresis in an ABI 3730xl sequencer (Applied Biosystems).

Factual Analysis

   Factual Data were examined by utilizing Epi Info rendition 3.2 ( Hazard factors for ESBL-creating E. coli were evaluated by utilizing univariate investigation with the χ2 or Fisher correct tests; notwithstanding, we utilized examination of change to decide if age had a relationship with ESBL-delivering E. coli. The importance edge was 0.05.


   Out of the 530 uropathogeneous E. coli confines, one hundred and forty seven (147) 27.7% multidrug safe E. coli , hypothetical ESBL test was done utilizing the DDST test, 47 (8.9%) was sure for ESBL, from the 47 positive ESBL, 14 conveyed CTX-M qualities from the forward and invert portrayal. The ESBL segregates demon - strated most elevated vulnerability to carbapenems and amikacin which is as per 2011 CLSI criteria for MIC test. The most astounding rate of protection was watched for the accompanying anti-infection agents: cefixime, colistin and ciprofloxacin.

Figure 1
Antibiotics Resistant (R%) Susceptible (S%)
Cefixime 100 (18.9%) 430 (81%)
Ceftriaxone 88 (16.6%) 442 (83.4%)
Ceftazidime 150 (20.3%) 380 (71.7%)
Cefotaxime 126 (23.8%) 404 (76%)
Augumentin 350 (66) 180 (34%)
Imipenem 47 (8.9%) 483 (91%)
Colistin 92 ( 17.3%) 438 (82.9%)
Ciprofloxacin 100 (18.9%) 430 (81%)
Amikacin 260 (49%) 270 (51%)

Table 1: Percentage antibiotics susceptibility profile across the isolates.
Key: R: Resistant; S: Susceptible; %: Percentage

Total Number of Isolate ESBL Positive ESBL Negative
147 47 (31.9%) 100 (68%)

Table 2: Showing the Double disk synergy test for the detection of ESBL across the multi drug resistant isolate.

Number of Isolate CTX m positive CTXm Negative
47 14 (29.7%) 33 ( 70%)

Table 3: Showing the reverse and forward characterization of the presumptive positive ESBL isolates.


   Relationship of Antimicrobial Resistance with Resistant Genes, demonstrates the nearness of safe qualities, and protection from various antimicrobial operators. As a rule, there were no notewor - thy contrasts in regards to nearness or nonappearance of qualities articulation. Strangely, higher protection from cefotaxime, amika- cin, and ceftriaxone was found in TEM negative. Protection from cotrimoxazole, imipenem, amikacin, and third era cephalosporins was watched more in CTX-M positive segregates than in CTX-M negative separates.

   In this investigation, an endeavor has been made to genotype the ESBL-creating E. coli disengages utilizing the forward and turn around attributes of the CTXm qualities related with multi tran- quilize safe Escherichia coli among pregnant moms introduced at the counter natal centers. The relationship of CTXm qualities with antimicrobial protection is seen with the expanding commonness of ESBL-creating E. coli (8.9%) and their abnormal state of protec - tion from expansive range anti-microbial agents.

   The predominance of ESBL-creating E. coli confines fluctuates in various parts of the world and even among various healing cen- ters inside a nation. The rate of predominance in our middle was around 8.9% which is near the outcomes announced by different examinations [11-25] and like rates of ESBL-delivering E. coli re - vealed in nations, for example, India (27%), Lebanon (13.3%), Ko - rea (9.2%), and turkey (17%) [26,27] .

   In, our examination the disengages demonstrated high protec- tion from amikacin, and colistin, which is additionally found in an investigation by Babypadmini and Appalaraju which revealed around 74% protection from trimethoprim/sulfamethoxazole and 91.6% protection from fluoroquinolone in ESBL-delivering E. coli pathogens by circle dispersion technique [29], which is con- siderably higher than our outcomes (65% protection from trime - thoprim/sulfamethoxazole and 76%. Results from different exam - inations from Malaysia and Spain demonstrated lower protection from trimethoprim/sulfamethoxazole and ciprofloxacin in pee tests from grown-ups than this investigation which might be be- cause of various patient populace (grown-ups versus pediatrics). Absolutely, the expanding protection of E. coli to trimethoprim makes this medication less powerful as empiric treatment of UTI [30-35].

  The protection from cefotaxime in CTX-M makers in our inves - tigation was higher than those revealed around 29.7% of CTX-M creating strains were impervious to quinolones and aminoglyco- sides, individually, which was higher contrasted with Edelstein., et al. [36] contemplate (21%) however it was lower than the Men - donca., et al. think about (93%).


  In spite of the fact that the CTXm was observed to be the second most predominant safe quality, the pervasiveness of other quali - ties alongside antimicrobial protection is on the ascent. Carbapen - ems were the best anti-infection agents against ESBL-delivering E. coli in urinary tract contamination among pregnant moms.

Conflict of Interest

There was no conflict of interest during and after this study


  We wish to extend our gratitude to the staff of health services department, University of Benin antenatal unit and all staff and members of Medical laboratory Science Department, University of Benin


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Copyright: © 2018 Otabor Florence Enogiomwan and Isaiah Nnanna Ibeh. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

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