Molecular Characterization of Multidrug-Resistant Escherichia coli Isolated from Milk of Dairy Cows with Clinical Mastitis in Algeria

ABSTRACT The objective of this study was to investigate the occurrence of multidrug-resistant Escherichia coli in cows with clinical mastitis in 42 different dairy farms located in the Bordj Bou Arreridj region of Algeria. Milk samples were cultured on Columbia blood agar, and isolates were then identified by matrix-assisted laser desorption ionization–time of flight mass spectrometry. In total, 200 samples were screened and 52 E. coli strains confirmed as causative agents were obtained. The antimicrobial susceptibility testing was performed by disk diffusion method. Antibiotic resistance genes, including those conferring resistance to extended-spectrum β-lactamases (i.e., blaTEM, blaSHV, and blaCTX-M), tetracyclines (tetA, tetB, tetC, and tetJ), aminoglycosides [aph(3′), aac(3′), aac(6′), ant, aad, and armA], and quinolones (qnrA and qnrB) were amplified by standard PCR and sequenced when positive. Transferability of resistance genes has been investigated by conjugation experiments and multilocus sequence typing. The most frequently observed resistance was to amoxicillin (86.5%), followed by tetracycline (75%), amoxicillin–clavulanic acid (59.6%), trimethoprim-sulfamethoxazole (36.5%), doxycycline (13.5%), and ciprofloxacin (13.5%). Multidrug resistance was observed in 38.4% of isolates. Genotypic characterization showed that tetA (44.2%) and blaTEM-1 (30.7%) genes were the most prevalent. Screening for plasmid-mediated quinolone resistance genes demonstrated that seven isolates (13.5%) expressed qnrB and one isolate (1.9%) harbored qnrA. In addition, aminoglycoside resistance determinants including aadA1 and aac(3)-Id were detected in seven and two isolates, respectively. Moreover, blaTEM, tetA, tetB, qnrB, and aadA1 were successfully transferred horizontally to transconjugant strains. The multilocus sequence typing revealed the presence of three different sequence types (ST162, ST371, and ST 949).

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Prevalence of class 1 and 2 integrons among the multidrug resistant uropathogenic strains of Escherichia coli

Asian Biomedicine ◽

10.5372/1905-7415.0901.367 ◽

2017 ◽

Vol 9 (1) ◽

pp. 49-54 ◽

Cited By ~ 1

Author(s):

Haddadi Azam ◽

Somayeh Mikaili Ghezeljeh ◽

Shavandi Mahmoud

Keyword(s):

Escherichia Coli ◽

Antibiotic Resistance ◽

Resistance Genes ◽

Antibiotic Resistance Genes ◽

Multidrug Resistant ◽

Diffusion Method ◽

Disk Diffusion Method ◽

Integrase Gene ◽

Class 1 ◽

Tract Infections

Abstract Background Multidrug resistance is a serious problem in the treatment of urinary tract infections. Horizontal gene transfer, directed by strong selective pressure of antibiotics, has resulted in the widespread distribution of multiple antibiotic resistance genes. The dissemination of resistance genes is enhanced when they are trapped in integrons. Objectives To determine the prevalence of integrons among multidrug resistant Escherichia coli strains collected from regional hospitals and private clinical laboratories in Alborz province. Methods The susceptibility of 111 clinical Escherichia coli isolates was tested using a Kirby–Bauer disk diffusion method for common antibiotics. Isolates were screened for the production of extended spectrum β-lactamases (ESBLs) using a double disk synergy test. The existence of integrons was confirmed by amplification of the integrase gene and their class determined via analysis of PCR products by PCR-RFLP. Results Isolates showed the highest resistance to amoxicillin. Nitrofurantoin, amikacin, and ceftizoxime were the most effective antibiotics in vitro. Eighty-eight isolates of 111 (79%) were resistant to more than three unrelated drugs. We found 30% of the multidrug resistant isolates harbor integrons. Class 1 and 2 integrons were detected in 25 and 1 isolates, respectively. ESBL screening of strains showed 45 isolates (40%) were positive; 22% of the ESBL-positive isolates carried class 1 integrons and the frequency of MDR in ESBLpositive isolates was 93%. Conclusion The existence of integrons in only 29.5% of multidrug resistant isolates showed that besides integrons, antibiotic resistance genes were probably carried on other transferable elements lacking integrons, such as transposons or plasmids.

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Molecular Characterization of β-Lactamase Producing Genes and Integrons in Diarrheagenic Escherichia Coli From Diarrheal Children Less Than Five Years of Age in Ouagadougou, Burkina Faso.

10.21203/rs.3.rs-628039/v1 ◽

2021 ◽

Author(s):

Rene Dembele ◽

Wendpoulomdé A.D. Kaboré ◽

Issiaka Soulama ◽

Oumar Traoré ◽

Nafissatou Ouédraogo ◽

...

Keyword(s):

Escherichia Coli ◽

Molecular Characterization ◽

Resistance Genes ◽

Antibiotic Resistance Genes ◽

Control Measures ◽

Diffusion Method ◽

Biochemical Tests ◽

Disk Diffusion Method ◽

E Coli

Abstract Background The aim of this study was to determine the resistance of diarrheagenic Escherichia coli strains to β-lactams antibiotics and to perform the molecular characterization of Extended Spectrum β-lactamases (ESBL) and integrons genes. Methods This study was carried out from August 2013 to October 2015 and involved 31 DEC strains isolated from diarrheal stools samples collected from children less than five years of age. The identification and characterization of DEC strains was done through the standard biochemical tests those were confirmed using API 20E and Polymerase Chain Reaction (PCR). The determination of antimicrobial resistance was realized by the disk diffusion method then an amplification of the β-lactamase resistance genes and integrons by PCR was done. Results Out of the 419 E. coli strains identified, 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 from the 31 DEC isolates: blaTEM (6.5%), blaSHV (19.4%), blaOXA (38.7%) blaCTX−M (9.7%), Int1 (58.1%) and Int3 (19.4%). No class 2 integrons (Int2) was characterized. Conclusions Because of the high prevalence of multidrug-resistant ESBL organisms found in this study among pediatric patients, there is a need of stringent pediatric infection control measures.

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Prevalence and Molecular Characterization of Antimicrobial Resistance in Escherichia coli Isolated from Raw Milk and Raw Milk Cheese in Egypt

Journal of Food Protection ◽

10.4315/0362-028x.jfp-17-277 ◽

2018 ◽

Vol 81 (2) ◽

pp. 226-232 ◽

Cited By ~ 8

Author(s):

Rabee A. Ombarak ◽

Atsushi Hinenoya ◽

Abdel-Rahman M. Elbagory ◽

Shinji Yamasaki

Keyword(s):

Escherichia Coli ◽

Antimicrobial Resistance ◽

Resistance Genes ◽

Raw Milk ◽

Diffusion Method ◽

Sequencing Analysis ◽

Aminoglycoside Resistance ◽

Disk Diffusion Method ◽

E Coli ◽

Extended Spectrum

ABSTRACT The goal of this study was to examine antimicrobial resistance and characterize the implicated genes in 222 isolates of Escherichia coli from 187 samples of raw milk and the two most popular cheeses in Egypt. E. coli isolates were tested for susceptibility to 12 antimicrobials by a disk diffusion method. Among the 222 E. coli isolates, 66 (29.7%) were resistant to one or more antimicrobials, and half of these resistant isolates showed a multidrug resistance phenotype (resistance to at least three different drug classes). The resistance traits were observed to tetracycline (27.5%), ampicillin (18.9%), streptomycin (18.5%), sulfamethoxazole-trimethoprim (11.3%), cefotaxime (4.5%), kanamycin (4.1%), ceftazidime (3.6%), chloramphenicol (2.3%), nalidixic acid (1.8%), and ciprofloxacin (1.4%). No resistance to fosfomycin and imipenem was observed. Tetracycline resistance genes tetA, tetB, and tetD were detected in 53 isolates, 9 isolates, and 1 isolate, respectively, but tetC was not detected. Aminoglycoside resistance genes strA, strB, aadA, and aphA1 were detected in 41, 41, 11, and 9 isolates, respectively. Sulfonamide resistance genes sul1, sul2, and sul3 were detected in 7, 25, and 3 isolates, respectively. Of 42 ampicillin-resistant isolates, blaTEM, blaCTX-M, and blaSHV were detected in 40, 9, and 3 isolates, respectively, and 10 (23.8%) ampicillin-resistant isolates were found to produce extended-spectrum β-lactamase. Each bla gene of extended-spectrum β-lactamase–producing E. coli was further subtyped to be blaCTX-M-15, blaCTX-M-104, blaTEM-1, and blaSHV-12. The class 1 integron was also detected in 28 resistant isolates, and three different patterns were obtained by PCR-restriction fragment length polymorphism. Sequencing analysis of the variable region revealed that four isolates had dfrA12/orfF/aadA2, two had aadA22, and one had dfrA1/aadA1. These data suggest that antimicrobial-resistant E. coli are widely distributed in the milk production and processing environment in Egypt and may play a role in dissemination of antimicrobial resistance to other pathogenic and commensal bacteria.

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Preliminary Results on the Prevalence of Salmonella Spp. in Marine Animals Stranded in Sicilian Coasts: Antibiotic Susceptibility Profile and ARGs Detection in the Isolated Strains

Pathogens ◽

10.3390/pathogens10080930 ◽

2021 ◽

Vol 10 (8) ◽

pp. 930

Author(s):

Delia Gambino ◽

Sonia Sciortino ◽

Sergio Migliore ◽

Lucia Galuppo ◽

Roberto Puleio ◽

...

Keyword(s):

Antibiotic Resistance ◽

Resistance Genes ◽

Antibiotic Susceptibility ◽

Antibiotic Resistance Genes ◽

Diffusion Method ◽

Salmonella Spp ◽

Marine Animals ◽

Disk Diffusion Method ◽

Phenotypic Resistance ◽

Low Prevalence

The presence of Salmonella spp. in marine animals is a consequence of contamination from terrestrial sources (human activities and animals). Bacteria present in marine environments, including Salmonella spp., can be antibiotic resistant or harbor resistance genes. In this study, Salmonella spp. detection was performed on 176 marine animals stranded in the Sicilian coasts (south Italy). Antibiotic susceptibility, by disk diffusion method and MIC determination, and antibiotic resistance genes, by molecular methods (PCR) of the Salmonella spp. strains, were evaluated. We isolated Salmonella spp. in three animals, though no pathological signs were detected. Our results showed a low prevalence of Salmonella spp. (1.7%) and a low incidence of phenotypic resistance in three Salmonella spp. strains isolated. Indeed, of the three strains, only Salmonella subsp. enterica serovar Typhimurium from S. coeruleoalba and M. mobular showed phenotypic resistance: the first to ampicillin, tetracycline, and sulphamethoxazole, while the latter only to sulphamethoxazole. However, all strains harbored resistance genes (blaTEM, blaOXA, tet(A), tet(D), tet(E), sulI, and sulII). Although the low prevalence of Salmonella spp. found in this study does not represent a relevant health issue, our data contribute to the collection of information on the spread of ARGs, elements involved in antibiotic resistance, now considered a zoonosis in a One Health approach.

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Molecular detection of antibiotic resistance genes in multidrug-resistant Staphylococcus aureus isolates from dog dental plaque

BULGARIAN JOURNAL OF VETERINARY MEDICINE ◽

10.15547/bjvm.2099 ◽

2019 ◽

Vol 22 (4) ◽

pp. 419-427

Author(s):

S. Nouri Gharajalar ◽

M. Onsori

Keyword(s):

Staphylococcus Aureus ◽

Antibiotic Resistance ◽

Multiplex Pcr ◽

Resistance Genes ◽

Dental Plaque ◽

Antibiotic Resistance Genes ◽

Multidrug Resistant ◽

Diffusion Method ◽

Pcr Assay ◽

Multiplex Pcr Assay

Multidrug resistant Staphylococcus aureus strains are a major health care problem both in humans and animals. In this work we described three multiplex PCR assays for detection of clinically relevant antibiotic resistance genes in S. aureus isolated from dog dental plaques. Thirty dental plaque samples were collected; then cultural, biochemical and molecular tests performed for isolation and identification of S. aureus from samples. The antibiotic susceptibility of the isolates were checked by Kirby Bauer disc diffusion method and the prevalence of antibiotic resistance genes determined using multiplex PCR assay. As a result S. aureus was isolated from 18 dog plaque samples. Fifteen of these isolates were resistant to penicillin. The mecA gene was more prevalent than blaZ among penicillin-resistant bacteria. Ten of the isolates were resistant to tetracycline. The percentage of tetM was higher than tetK among them. Also, 10 of the isolates were resistant to cefazolin among them bla TEM detected in higher rate than blaSHV and blaOXA-1. Hence multiplex PCR assay is a suitable method for detection of antibiotic resistance patterns of S. aureus isolates.

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Occurrence of NDM-1 and VIM-2 Co-Producing Escherichia coli and OprD Alteration in Pseudomonas aeruginosa Isolated from Hospital Environment Samples in Northwestern Tunisia

Diagnostics ◽

10.3390/diagnostics11091617 ◽

2021 ◽

Vol 11 (9) ◽

pp. 1617

Author(s):

Raouaa Maaroufi ◽

Olfa Dziri ◽

Linda Hadjadj ◽

Seydina M. Diene ◽

Jean-Marc Rolain ◽

...

Keyword(s):

Escherichia Coli ◽

Pseudomonas Aeruginosa ◽

Bacterial Resistance ◽

Carbapenem Resistance ◽

Multidrug Resistant ◽

Hospital Environment ◽

Diffusion Method ◽

Resistant Bacteria ◽

Multidrug Resistant Bacteria ◽

Disk Diffusion Method

Hospital environments constitute the main reservoir of multidrug-resistant bacteria. In this study we aimed to investigate the presence of Gram-negative bacteria in one Northwestern Tunisian hospital environment, and characterize the genes involved in bacterial resistance. A total of 152 environmental isolates were collected from various surfaces and isolated using MacConkey medium supplemented with cefotaxime or imipenem, with 81 fermenter bacteria (27 Escherichia coli, and 54 Enterobacter spp., including 46 Enterobacter cloacae), and 71 non-fermenting bacteria (69 Pseudomonas spp., including 54 Pseudomonas aeruginosa, and 2 Stenotrophomonas maltophilia) being identified by the MALDI-TOF-MS method. Antibiotic susceptibility testing was performed by disk diffusion method and E-Test was used to determine MICs for imipenem. Several genes implicated in beta-lactams resistance were characterized by PCR and sequencing. Carbapenem resistance was detected among 12 isolates; nine E. coli (blaNDM-1 (n = 8); blaNDM-1 + blaVIM-2 (n = 1)) and three P. aeruginosa were carbapenem-resistant by loss of OprD porin. The whole-genome sequencing of P. aeruginosa 97H was determined using Illumina MiSeq sequencer, typed ST285, and harbored blaOXA-494. Other genes were also detected, notably blaTEM (n = 23), blaCTX-M-1 (n = 10) and blaCTX-M-9 (n = 6). These new epidemiological data imposed new surveillance strategies and strict hygiene rules to decrease the spread of multidrug-resistant bacteria in this area.

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The Dissemination of Multidrug Resistance (MDR) and Extensively Drug Resistant (XDR) among Uropathogenic E. coli (UPEC) Isolates from Urinary Tract Infection Patients in Babylon Province, Iraq

Baghdad Science Journal ◽

10.21123/bsj.2019.16.4(suppl.).0986 ◽

2019 ◽

Vol 16 (4(Suppl.)) ◽

pp. 0986

Author(s):

Al-Hasnawy Et al.

Keyword(s):

Escherichia Coli ◽

Urinary Tract Infection ◽

Urinary Tract ◽

Tract Infection ◽

Multidrug Resistant ◽

Diffusion Method ◽

Drug Resistant ◽

Disk Diffusion Method ◽

E Coli ◽

Extensively Drug Resistant

Antibiotic resistance is a problem of deep scientific concern both in hospital and community settings. Rapid detection in clinical laboratories is essential for the judicious recognition of antimicrobial resistant organisms. So, the growth of Uropathgenic Escherichia coli (UPEC) isolates with Multidrug-resistant (MDR) and Extensively Drug-resistant (XDR) profiles that thwart therapy for (UTIs) has been detected and has straight squeezed costs and extended hospital stays. This study aims to detect MDR- and XDR-UPEC isolates. Out of 42 UPEC clinical isolates were composed from UTI patients. The bacterial strains were recognized by standard laboratory protocols. Susceptibility to antibiotic was measured by the standard disk diffusion method Out of 42 Uropathogenic E. coli, 37 (88.09%) were found to be MDR while 5 isolates (11.90%) were XDR. The present study concluded high prevalence of uropathogenic Escherichia coli (UPEC) with Multidrug-resistant (MDR) isolated from urinary tract infection in Babylon province – Iraq.

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Antimicrobial Resistance and Virulence Determination of Enterococcus spp. Obtained from Hospital Environment in Iran

10.21203/rs.3.rs-123398/v1 ◽

2020 ◽

Author(s):

Saba Asgharzadeh Marghmalek ◽

Reza Valadan ◽

Mehrdad Gholami ◽

Mohtaram Nasrolahei ◽

Hamid Reza Goli

Keyword(s):

Antimicrobial Resistance ◽

Resistance Genes ◽

Pathogenic Bacteria ◽

Antibiotic Resistance Genes ◽

Hospital Environment ◽

Diffusion Method ◽

Virulence Determinants ◽

Environmental Isolates ◽

Disk Diffusion Method ◽

Antimicrobial Resistance Genes

Abstract Background: The role of the hospital environment as a source of pathogenic bacteria in recent studies has been poorly investigated. This study investigated the distribution of antimicrobial resistance genes and virulence determinants in Enterococcus species isolated from hospital environment in Sari, Iran. Method: Overall, 90 enterococci strains were obtained from high touch surfaces of four hospitals in Sari, Iran. These environmental samples were obtained from bathroom, beds, tables, doorknobs, room keys, wheelchair and walls in the patient and staff’s rooms. The resistance profile of the isolates was determined by disk diffusion method. Seven resistance genes and two virulence associated genes were evaluated molecularly by multiplex PCR. Results: According to the PCR, 42 (46.66%) of them were E. faecalis and 48 (53.33%) others were detected as E. faecium. Also, 28 (66.6%) E. faecalis and 18 (37.5%) E. faecium isolates were multidrug-resistant (MDR). Among all 90 environmental isolates 54 (60%), 54 (60%), 8 (8.8%), 8 (8.8%), 60 (66.6%), 26 (28.8%), and 24 (26.6%) isolates contained tetM, tetL, vanA, vanB, ermB, aac(6´)-Ie-aph(2´´)-Ia, and aph (3´)-IIIa, respectively. Moreover, all isolates were investigated for the presence of virulence genes and 88 (97.7%) of isolates had esp gene, and 16 (17.7%) had ace.Conclusions: This report showed that the environmental isolates of Enterococcus are the major sources of antibiotic resistance genes that can transfer them to the clinical isolates of bacteria in hospital settings. An effective following strategy should be organized to clearance and stop emergence of these pathogenic bacteria.

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Biocide's susceptibility and frequency of biocide resistance genes and the effect of exposure to sub-inhibitory concentrations of sodium hypochlorite on antibiotic susceptibility of Stenotrophomonas maltophilia

10.1101/2021.10.04.463145 ◽

2021 ◽

Author(s):

Safar Ali Alizadeh ◽

Amir Javadi ◽

Farhad Nikkhahi ◽

Mohammad Rostamani ◽

Mehdi Bakht ◽

...

Keyword(s):

Sodium Hypochlorite ◽

Resistance Genes ◽

Antibiotic Susceptibility ◽

Inhibitory Concentration ◽

Microtiter Plate ◽

Multidrug Resistant ◽

Diffusion Method ◽

Dilution Method ◽

Disk Diffusion Method ◽

Microtiter Plate Assay

Background: The overused of biocides in healthcare-facilities poses risk for emergence and spread of antibiotic resistance among nosocomial pathogens. Hospital-acquired infections due to S. maltophilia particularly in the immunocompromised patients have been increased. The objective of this study was to evaluate the susceptibility of S. maltophilia clinical isolates to commonly used biocides in hospitals, as well the frequency of biocides resistance gene among them. This study also intended to assess the effect of exposure of S. maltophilia isolates to sub-inhibitory concentrations of sodium hypochlorite upon the antimicrobial susceptibility patterns. Methods: This study included 97 S. maltophilia isolates. Biofilm formation was determined by microtiter plate assay. The susceptibility tests of five biocides were studied against all S. maltophilia isolates by microbroth dilution method. Susceptibility of isolates to antibiotics by disk diffusion method were compared before and after exposure to sub-inhibitory concentrations of sodium hypochlorite. Presence of qacE, qacEΔ1, SugE genes was screened by PCR. Results: Based on minimum inhibitory and bactericidal concentrations of biocides sodium hypochlorite 5% and ethyl alcohol 70% were the strongest and weakest against S. maltophilia isolates, respectively. The frequency of sugE gene resistance genes was found to be high (90.7%) in our clinical S. maltophilia isolates. None of the isolates carried qacE and qacEΔ1 gene. Exposure to sub-inhibitory concentration of sodium hypochlorite showed significantly change the susceptibility of isolates towards ceftazidime (P = .019), ticarcillin/clavulanate (P = .009). and chloramphenicol (P = .028). Conclusions: This study demonstrated that exposure to sub-inhibitory concentration of sodium hypochlorite leads to reduced antibiotic susceptibility and development of multidrug-resistant S. maltophilia strains.

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Carbapenemases and Extended-Spectrum β-Lactamase–Producing Multidrug-Resistant Escherichia coli Isolated from Retail Chicken in Peshawar: First Report from Pakistan

Journal of Food Protection ◽

10.4315/0362-028x.jfp-18-045 ◽

2018 ◽

Vol 81 (8) ◽

pp. 1339-1345 ◽

Cited By ~ 2

Author(s):

KAFEEL AHMAD ◽

FARYAL KHATTAK ◽

AMJAD ALI ◽

SHAISTA RAHAT ◽

SHAZIA NOOR ◽

...

Keyword(s):

Escherichia Coli ◽

Antibiotic Resistance ◽

Nalidixic Acid ◽

Resistance Genes ◽

Antibiotic Resistance Genes ◽

Multidrug Resistant ◽

E Coli ◽

Extended Spectrum ◽

Amoxicillin Clavulanate ◽

Synergy Test

ABSTRACT We report the prevalence of extended-spectrum β-lactamases and carbapenemases in Escherichia coli isolated from retail chicken in Peshawar, Pakistan. One hundred E. coli isolates were recovered from retail chicken. Antibiotic susceptibility testing was carried out against ampicillin, chloramphenicol, kanamycin, nalidixic acid, cephalothin, gentamicin, sulfamethoxazole-trimethoprim, and streptomycin. Phenotypic detection of β-lactamase production was analyzed through double disc synergy test using the antibiotics amoxicillin-clavulanate, cefotaxime, ceftazidime, cefepime, and aztreonam. Fifty multidrug-resistant isolates were screened for detection of sul1, aadA, cmlA, int, blaTEM, blaSHV, blaCTX-M, blaOXA-10, blaVIM, blaIMP, and blaNDM-1 genes. Resistance to ampicillin, nalidixic acid, kanamycin, streptomycin, cephalothin, sulfamethoxazole-trimethoprim, gentamicin, cefotaxime, ceftazidime, aztreonam, cefepime, amoxicillin-clavulanate, and chloramphenicol was 92, 91, 84, 73, 70, 67, 53, 48, 40, 39, 37, 36, and 23% respectively. Prevalence of sul1, aadA, cmlA, int, blaTEM, blaCTX-M, blaIMP, and blaNDM-1 was 78% (n = 39), 76% (n = 38), 20% (n = 10), 90% (n = 45), 74% (n = 37), 94% (n = 47), 22% (n = 11), and 4% (n = 2), respectively. blaSHV, blaOXA-10, and blaVIM were not detected. The coexistence of multiple antibiotic resistance genes in multidrug-resistant strains of E. coli is alarming. Hence, robust surveillance strategies should be developed with a focus on controlling the spread of antibiotic resistance genes via the food chain.

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