Caracterización de Escherichia coli D7111 productora de ß-LACTAMASA TEM-176 / Characterization of Escherichia coli D7111 producing the ß-LACTAMASA TEM-176

RESUMEN El presente reporte es la descripción original de bla TEM-176. Se caracterizaron los mecanismos de resistencia a antimicrobianos de un aislamiento de Escherichia coli enterotoxigénica, determinándose la resistencia a 22 antimicrobianos categorizados en 15 grupos diferentes mediante difusión en agar, estableciéndose grupo filogenético, mecanismos de resistencia y presencia de integrones de Clase 1 y 2 mediante PCR. Integrones y genes de resistencia a β-lactámicos fueron secuenciados. El aislamiento del grupo filogenético A, mostró resistencia o sensibilidad disminuida a ampicilina, amoxicilina más ácido clavulánico, ácido nalidíxico, ciprofloxacino, estreptomicina, kanamicina, tetraciclina, trimetoprim, sulfisoxazol, cotrimoxazol, azitromicina y nitrofurantoina, detectándose la presencia de bla TEM, aadA1/2, aphA1, sul3, tet(A) y un integron de Clase 2 conteniendo un gen dfrA1. La resistencia a quinolonas se relacionó con la substitución Ser83Ala. La secuencia de TEM mostró la substitución Ala222Val, la cual a la fecha no había sido descrita, reportándose como una nueva β-lactamasa, con el nombre de bla TEM-176.

ABSTRACT The present report is the original description of bla TEM-176. The mechanisms of resistance to antimicrobial agents were determined in an enterotoxigenic Escherichia coli, determining the susceptibility to 22 antimicrobials classified in 15 different groups by agar diffusion and establishing the phylogenetic group, mechanisms of resistance and presence of Class 1 and 2 integrons. Integrons and β-lactam resistance genes were sequenced. The isolate, belonging to phylogenetic group A, showed the presence of resistance or diminished susceptibility to a ampicillin, amoxicillin plus clavulanic acid, nalidíxic acid, ciprofloxacin, streptomycin, kanamycin, tetracycline, trimethoprim, sulfisoxazole, cotrimoxazole, azithromycin and nitrofurantoin, carrying bla TEM, aadA1/2, aphA1, sul3, tet(A) and a Class 2 integron containing a dfrA1 gene. Quinolone resistance was related to the substitution Ser83Ala. The TEM sequencing showed the presence of the new substitution Ala222Val, which led to the description of the new β-lactamase bla TEM-176.

Caracterización de Escherichia coli D7111 productora de ß-LACTAMASA TEM-176 / Characterization of Escherichia coli D7111 producing the ß-LACTAMASE TEM-176

RESUMEN El presente reporte es la descripción original de bla TEM-176. Se caracterizaron los mecanismos de resistencia a antimicrobianos de un aislamiento de Escherichia coli enterotoxigénica, determinándose la resistencia a 22 antimicrobianos categorizados en 15 grupos diferentes mediante difusión en agar, estableciéndose grupo filogenético, mecanismos de resistencia y presencia de integrones de Clase 1 y 2 mediante PCR. Integrones y genes de resistencia a β-lactámicos fueron secuenciados. El aislamiento del grupo filogenético A, mostró resistencia o sensibilidad disminuida a ampicilina, amoxicilina más ácido clavulánico, ácido nalidíxico, ciprofloxacino, estreptomicina, kanamicina, tetraciclina, trimetoprim, sulfisoxazol, cotrimoxazol, azitromicina y nitrofurantoina, detectándose la presencia de bla TEM, aadA1/2, aphA1, sul3, tet(A) y un integron de Clase 2 conteniendo un gen dfrA1. La resistencia a quinolonas se relacionó con la substitución Ser83Ala. La secuencia de TEM mostró la substitución Ala222Val, la cual a la fecha no había sido descrita, reportándose como una nueva β-lactamasa, con el nombre de bla TEM-176.

ABSTRACT The present report is the original description of bla TEM-176. The mechanisms of resistance to antimicrobial agents were determined in an enterotoxigenic Escherichia coli, determining the susceptibility to 22 antimicrobials classified in 15 different groups by agar diffusion and establishing the phylogenetic group, mechanisms of resistance and presence of Class 1 and 2 integrons. Integrons and β-lactam resistance genes were sequenced. The isolate, belonging to phylogenetic group A, showed the presence of resistance or diminished susceptibility to a ampicillin, amoxicillin plus clavulanic acid, nalidíxic acid, ciprofloxacin, streptomycin, kanamycin, tetracycline, trimethoprim, sulfisoxazole, cotrimoxazole, azithromycin and nitrofurantoin, carrying bla TEM, aadA1/2, aphA1, sul3, tet(A) and a Class 2 integron containing a dfrA1 gene. Quinolone resistance was related to the substitution Ser83Ala. The TEM sequencing showed the presence of the new substitution Ala222Val, which led to the description of the new β-lactamase bla TEM-176.

Antimicrobial resistance patterns and prevalence of integrons in Shigella species isolated from children with diarrhea in southwest Iran

Objective: To investigate the antimicrobial resistance patterns and prevalence of integrons in Shigella species isolated from children with diarrhea in southwest Iran. Methods: In this study, 1 530 stool samples were collected from children under 15 years with diarrhea referred to teaching hospitals in Ahvaz and Abadan, southwest Iran. Shigella spp. were identified by standard biochemical tests and PCR. The antibiotic resistance pattern of all Shigella isolates was determined by the disk diffusion method and minimum inhibitory concentration (MIC) by E-test. Results: Of 1 530 stool samples, 91 (5.9%, 91/1 530) were positive for Shigella spp. the most common Shigella isolates were Shigella flexneri 47 (51.6%, 47/1 530). Antibiotic susceptibility tests showed that the highest antibiotic resistance was related to trimethoprim-sulfamethoxazole (87.9%, 80/91) and ampicillin (86.8%, 79/91). Multiplex PCR results revealed that 56% and 86.9% of Shigella isolates carried integron class I and integron class II genes, respectively. None of the isolates included the integron class III gene. Conclusions: The high prevalence of multi-drug resistance in Shigella isolates in our area increases the concerns about dissemination of the antibiotic-resistant isolates in this bacterium.

Corynebacterium pseudotuberculosis biovar ovis: evaluación de la sensibilidad antibiótica in vitro / Corynebacterium pseudotuberculosis biovar ovis: Evaluation of antibiotics susceptibility in vitro

Resumen Los objetivos de este trabajo fueron estudiar la sensibilidad antibiótica de aislamientos de Corynebacterium pseudotuberculosis procedentes de pequeños rumiantes e investigar la presencia de integrones que contienen genes de resistencia. Se estudiaron 15 aislamientos de diferentes fuentes por los métodos de difusión y dilución. Por el método de difusión, amoxicilina-clavulánico, ampicilina, cefotaxima, cefoxitina, ciprofloxacina, cloranfenicol, eritromicina, estreptomicina, gentamicina, imipenem, kanamicina, norfloxacina, penicilina, rifampicina, tetraciclina, trimetroprima-sulfametoxazol y vancomicina fueron activos frente al 100% de los aislamientos, mientras que amicacina presentó resultados variables. En los aislamientos que desarrollaron frente a amicacina se investigó la presencia de integrones de clase 1. El resultado fue negativo, sugiriendo la ausencia del integrón. Utilizando el método de dilución, los antibióticos más activos correspondieron a los grupos de cefalosporinas, gluco-péptidos, macrólidos, quinolonas y tetraciclinas. Se demostró menor actividad de p-lactámicos y aminoglucósidos. No se registró variabilidad en los perfiles antibióticos en los aislamientos procedentes de diferentes fuentes.

Abstract The aims of this work were to study the antibiotic susceptibility in Corynebacterium pseudotuberculosis isolated from small ruminants and to determine the presence of integrons that contain resistance genes. Fifteen isolates of different sources were analysed using the diffusion and the dilution methods. When the diffusion method was performed, amoxicillin-clavulanic, ampicillin, cefotaxime, cefoxitin, ciprofloxacin, chloramphenicol, erythromycin, streptomycin, gentamicin, imipenem, kanamycin, norfloxacin, penicillin, rifampicin, tetracycline, trimethoprim-sulfamethoxazole and vancomycin were effective against the 100% of isolates, while amikacin showed variable results. The isolates that were able to grow with amikacin, were studied in relation to the presence of integron class 1. The result was negative, suggesting the absence of integron. Using dilution method, the antibiotics belonging to the cephalosporin, glycopeptide, macrolide, quinolone, and tetracycline groups were the most active ones for the C. pseudotuberculosis biovar ovis isolates. Less activity of p-lactam and aminoglycosides were observed. There was no observation of variability in the antibiotic patterns in the strains coming from different sources.

Perfil de resistência aos antibióticos e prevalência dos genes qacEΔ1 e sul1 em Pseudomonas aeruginosa de efluente hospitalar / Antibiotic resistance pattern and prevalence of qacEΔ1 and sul1 genes in Pseudomonas aeruginosa from hospital wastewater

Introdução: Efluentes hospitalares representam riscos à saúde pública e ambiental devido à presença de microrganismos patogênicos, drogas e produtos químicos. Pseudomonas aeruginosa é um patógeno oportunista frequentemente encontrado no ambiente hospitalar. Objetivo: Avaliar o resistoma de isolados de P. aeruginosa da estação de tratamento de esgoto hospitalar (ETEH) de um complexo hospitalar na cidade do Rio de Janeiro. Método: Vinte isolados dos cinco estágios da ETEH foram identificados como P. aeruginosa pelo sequenciamento do gene 16S rRNA. A suscetibilidade aos antibióticos foi determinada segundo o CLSI e os genes qacEΔ1 e sul1 foram detectados pela PCR. Resíduos de sulfonamidas foram pesquisados por cromatografia líquida de alta eficiência acoplada à espectrometria de massas sequencial. Resultados: Foi demonstrada a presença de sulfametoxazol em nível inferior a 50 ng∙L−1, resistência às sulfonamidas (80%) seguida pelas quinolonas (50%) e 13 perfis de suscetibilidade aos antimicrobianos. Os genes qacEΔ1-sul1 foram detectados em 100% dos isolados, sugerindo a presença de integrons de classe 1 em toda a ETEH. Conclusões: Os resultados sinalizaram limitações no tratamento e a propagação de genes de resistência nas etapas da ETEH. Esses dados contribuem com órgãos competentes no desenho de ações preventivas frente aos impactos negativos à saúde pública.

Introduction: Hospital effluents may pose great environmental risk due to the presence of pathogenic microorganisms, drugs and chemical components. Pseudomonas aeruginosa is an opportunistic pathogen frequently found in hospital environment. Objective: To evaluate the resistome of P. aeruginosa from the hospital wastewater treatment plant (HWTP) in a hospital complex of Rio de Janeiro city. Method: Twenty isolates from the five stages of the HWTP were identified as P. aeruginosa by 16S rRNA gene sequencing analysis. Susceptibility to antibiotics was determined according to CLSI and qacEΔ1 and sul1 genes were detected by PCR. Sulphonamide residues were investigated by high performance liquid chromatography coupled to sequential mass spectrometry. Results: The sulfamethoxazole has been demonstrated at a level below 50 ng L-1. Sulfonamide resistance (80%) has been demonstrated followed by quinolone class (50%) and 13 susceptibility patterns to antimicrobials. The qacEΔ1-sul1 genes were detected in 100% of isolates suggesting the presence of class 1 integrons in the whole HWTP. Conclusions: The results signalized limitations of HWTP and propagation of resistance genes in all stages of the HWTP. These data also contribute to the environmental sanitary surveillance in the design of prevention actions against negative impact on the public health.

Changing facades of Vibrio cholerae: An enigma in the epidemiology of cholera

Cholera, caused by the Gram-negative bacterium Vibrio cholerae, has ravaged humanity from time immemorial. Although the disease can be treated using antibiotics along with administration of oral rehydration salts and controlled by good sanitation, cholera is known to have produced mayhems in ancient times when little was known about the pathogen. By the 21st century, ample information about the pathogen, its epidemiology, genetics, treatment and control strategies was revealed. However, there is still fear of cholera outbreaks in developing countries, especially in the wake of natural calamities. Studies have proved that the bacterium is mutating and evolving, out-competing all our efforts to treat the disease with previously used antibiotics and control with existing vaccines. In this review, the major scientific insights of cholera research are discussed. Considering the important role of biofilm formation in the V. cholerae life cycle, the vast availability of next-generation sequencing data of the pathogen and multi-omic approach, the review thrusts on the identification of suitable biofilm-inhibiting targets and the discovery of anti-biofilm drugs from nature to control the disease.

The presence of plasmids in Aeromonas hydrophila and its relationship with antimicrobial and heavy metal-resistance profiles / Presença de plasmídeos em Aeromonas hydrophila e sua relação com perfis de resistência aos antimicrobianos e metais pesados

ABSTRACT: Aeromonas hydrophila is a common fish pathogen that causes extensive damage to aquaculture. To develop and implement a more adequate strategy to farm fish, it is crucial to understand the bacterial-resistance levels and their transference dynamics. The objective of this study was to analyze the resistance profile of isolated Aeromonas hydrophila to antimicrobial agents and heavy metals and draw a correlation of the observed profiles with the presence of plasmids. Resistance of the isolated bacteria to antimicrobial agents (oxacilin, gentamicin, tetracycline, and nalidixic acid) and heavy metals (cadmium, lead, copper, and manganese) was verified using the minimum bactericidal concentration (MBC) and minimum inhibitory concentration (MIC) standards. The Multiple Antibiotic Resistance Index (MAR Index) was calculated. Plasmids were extracted by using a common methodology described elsewhere. Mann-Whitney Test, implemented in the R environment, was used to determine the correlation between resistance and plasmids presence. A high resistance to almost all antimicrobial agents and heavy metals was observed, except to gentamicin and cadmium. The MAR index results showed resistance to all antimicrobial profiles. Of the isolated bacteria, 14 showed the presence of plasmids. However, no correlation was noted between the resistance profile and the plasmid presence for these isolates, indicating that the genes responsible for resistance to microbial agents and heavy metals are present in the cromossomic DNA, which in turn suggested the possibility of gene transfer between the isolated bacteria. The resistance to heavy metals can be linked to heavy utilization of fertilizers along the Sao Francisco River.

RESUMO: Bactérias da espécie Aeromonas hydrophila são patógenos que atacam peixes, causando grandes prejuízos à piscicultura. Entender os perfis de resistência dessa bactéria e a capacidade da mesma em transferir tal resistência é importante para implantação de um manejo adequado na produção de peixes. Os objetivos desse estudo foram analisar a resistência de isolados de Aeromonas hydrophila à antimicrobianos e metais pesados e, correlacionar os perfis encontrados com a presença de plasmídeos. A resistência dos isolados aos antimicrobianos (oxacilina, gentamicina, tetraciclina e ácido nalidíxico) e metais pesados (cádmio, chumbo, cobre e manganês) foi verificada pelas técnicas da Concentração Bactericida Mínima (CBM) e Concentração Inibitória Mínima (CIM). Foi calculado o Índice de Resistência Múltipla aos Antimicrobianos (IRMA). Os plasmídeos foram extraídos por metodologias descritas pela literatura. A relação entre a resistência aos antimicrobianos e metais pesados com a presença de plasmídeos foi determinada pelo teste de Mann-Whitney utilizando o ambiente R. Foi observada alta resistência aos antimicrobianos e metais pesados testados, com exceção à gentamicina e cádmio. No IRMA os isolados apresentaram resistência a todos os perfis de antimicrobianos possíveis. Quatorze isolados apresentaram plasmídeos, mas não foi encontrada relação dos perfis de resistência com a presença destes, o que indica que os genes de resistência a esses compostos estejam presentes no DNA cromossômico. Porém apontam a possibilidade de transferência dos genes de resistência entre os isolados. Estes resultados apontam alta resistência dos isolados e capacidade de transmissão dessa resistência a outras bactérias. A resistência aos metais pesados, pode estar ligada ao uso de fertilizantes nas plantações localizadas próximas as margens do Rio São Francisco.

Influence of antibiotic concentration on the integration frequency of resistance cassette in the integron / 中华检验医学杂志

Objective@#To observe the integration frequency of aadA2 resistance cassette at attI site of the integron under different concentration of streptomycin. @*Methods@#Class 1 integron with known gene sequence was cloned into plasmid pACYC184 to produce recombinant plasmid pACIDA, meanwhile the integrase gene was cloned into plasmid pET28a to construct recombinant plasmid pETINT. These two recombinant plasmids were consecutively transformed into E. coli BL(DE3). These transformed bacteria was cultured in the LB medium at 37 ℃ overnight with addition of different concentration of streptomycin. The copy number of total integrons and the copy number of integrated aadA2 at attI site of integrons were determined by using real-time PCR. and the integration frequency is the result of the former divided by the latter. @*Results@#The resulting frequencies were (1.97±0.24)×10-3, (3.23±1.77)×10-3, (3.27±0.67)×10-3, 0.45±0.13 and 1.32±0.11, with respective streptomycin concentrations of 0, 20, 30, 40 and 50 μg/ml. The background frequency of integration without integrase overexpression was less than (1.75±0.33)×10-7. @*Conclusion@#These findings indicate that antibiotic concentration significantly increase recombination frequency of aadA2 resistance cassette at attI site of the integron, catalyzed by integron integrase.(Chin J Lab Med, 2018, 41: 531-535)

Molecular detection of ß-lactamase and integron genes in clinical strains of Klebsiella pneumoniae by multiplex polymerase chain reaction

Abstract INTRODUCTION: Infections caused by β-lactamase-producing gram-negative bacteria, such as Klebsiella pneumoniae, are increasing globally with high morbidity and mortality. The aim of the current study was to determine antimicrobial susceptibility patterns and the prevalence of antibiotic resistance genes (β-lactamase and integron genes) using multiplex PCR. METHODS One-hundred K. pneumoniae isolates were collected from different clinical samples. Antibiotic susceptibility testing was performed with thirteen different antibiotics. Multiplex-PCR was used to detect β-lactamase (bla TEM, bla CTX-M, bla SHV , bla VEB, bla PER, bla GES, bla VIM, bla IMP, bla OXA, and bla KPC) and integron genes (int I, int II, and int III). RESULTS: The highest and lowest rate of resistance was exhibited against amikacin (93%) and imipenem (8%), respectively. The frequency of β-lactamase-positive K. pneumoniae was 37%, and the prevalence of the bla TEM, bla CTX-M, bla SHV , bla VEB, bla PER, bla GES, bla VIM, bla IMP, bla OXA, and bla KPC genes was 38%, 24%, 19%, 12%, 6%, 11%, 33%, 0%, 28%, and 23%, respectively. Of the 100 isolates, eight (8%) were positive for class I integrons; however, class II and III integrons were not detected in any of the strains. CONCLUSIONS: These results indicate co-carriage of a number of β-lactamase genes and antibiotic resistance integrons on the same plasmids harboring multi-drug resistance genes. It seems that these properties help to decrease treatment complications due to resistant bacterial infections by rapid detection, infection-control programs and prevention of transmission of drug resistance.

Differentiation of Pc and P2 variants in class 1 integron by high-resolution melting analysis / 中华检验医学杂志

Objective To develop a simple high-resolution melting ( HRM) analysis method for differentiation of Pc and P2 variants in class 1 integron.Methods DNA fragments containing Pc and P2 variants were amplified from plasmids pACW ( PcW ) and pACWP2 ( PcW-P2 ) respectively , then these purified PCR products and P 2 promoters were analyed full-length amplicon by HRM .Eight DNA fragments containing different Pc promoters were amplified and site-specific mutated from plasmids pACS ( PcS ) , pACH2 ( PcH2 ) , pACH1 ( PcH1 ) , pACW ( PcW ) , genomic DNA of Klebsiellar pneumonia HS07-68 (PcWTGN-10)and HS05-1792(PcH2TGN-10)respectively.The purified PCR products and eight Pc variants were characterized by HRM analyses of an unlabeled probe and full-length amplicon.This assay was applied to the differentiate Pc and P2 variants in 109 class 1 integrons from 95 urine clinical Escherichia coli isolates in Huashan Hospital during 2004 -2007.The differentiation results were compared with that determined by direct sequencing .Results P2 promoter with a significant higher melting temperature ( Tm ) can be identified by HRM analysis clearly .P2 promoters were identified in 2 class 1 integrons and consistent with direct sequencing results .Eight Pc variants were classified into three groups: PcS, PcSTGN-10 , PcW, PcWTGN-10, PcH1, PcH1TGN-10.Using direct HRM analysis.PcH2, PcH2TGN-10 were classified into four groups:PcS, PcH1, PcH2, PcW, PcSTGN-10 , PcH1TGN-10 , PcH2TGN-10 , PcWTGN-10 according to the melting curves of the unlabeled probe .Combined the HRM analyses of the whole amplicon and unlabeled probe , the eight Pc variants can be differentiated from each other .Five different Pc variants, PcS, PcW, PcH1, PcH2TGN-10 and PcWTGN-10 , were identified and consistent with direct sequencing results .Conclusions This developed a simple Pc and P 2 variants differentiation method via simultaneous HRM analyses of an unlabeled probe and full-length amplicon .This method is cost-effective and accurate , could be used in differentiation of Pc and P2 variants of class 1 integrons in clinical isolates .

Progress in the research of integron in bacterial resistance / 中华检验医学杂志

Integron was a novel bacterial resistance gene horizontal transmission element.In recent years, many researchers made a lot of research on the resistance mechanism of pathogen.In this paper, the detection and the novel discovery of the integron gene cassette were summarized , including Enterobacteria, Acinetobacter baumanii, Pseudomonas aeruginosa and other bacteria , on the basis of the multidrug resistance mechanism mediated by integron.The prospect the research was descixbed and more attention should be paid to the prevention and control of multidrug resistant bacteria.

Detection of Integrons and Staphylococcal Cassette Chromosome mec Types in Clinical Methicillin-resistant Coagulase Negative Staphylococci Strains

OBJECTIVES: Integrons are thought to play an important role in the spread of antibiotic resistance. This study investigates class 1 and 2 integron-positive methicillin-resistant coagulase-negative staphylococci strains isolated in Iran and characterizes their patterns of antimicrobial resistance. METHODS: Hundred clinical isolates of coagulase-negative staphylococci were characterized for integron content and staphylococcal cassette chromosome mec (SCCmec) type. RESULTS: Sixteen isolates carried class 1 (intI1) integrons and four isolates carried class 2 (intI2) integrons. One resistance gene array was identified among the class 1 integrons (aadA1 cassette). The distribution of SCCmec types in 50 methicillin-resistant coagulase-negative staphylococci strains showed that SCCmec types III and V dominated among the tested strains. CONCLUSION: This is the first report of methicillin-resistant coagulase-negative staphylococci strains that carry two mobile genetic elements, including class 1 and 2 integrons and SCCmec, in Iran.

Phenotypic and genotypic antimicrobial resistance and virulence genes of Salmonella enterica isolated from pet dogs and cats

Salmonella enterica isolates (n = 122), including 32 serotypes from 113 dogs and 9 cats, were obtained from household dogs (n = 250) and cats (n = 50) during 2012–2015. The isolates were characterized by serotyping, antimicrobial resistance phenotyping and genotyping, and virulence gene screening. Serovars Weltevreden (15.6%) and Typhimurium (13.9%) were the most common. The majority (43%) of the isolates were multidrug resistant. The dog isolates (12.3%) harbored class 1 integrons, of which the dfrA12-aadA2 cassette was most frequent (66.7%). The only class integron in serovar Albany was located on a conjugative plasmid. Two ESBL-producing isolates (i.e., a serovar Krefeld and a serovar Enteritridis) carried bla(TEM) and bla(CTX-M), and the bla(TEM) gene in both was horizontally transferred. Of the plasmid-mediated quinolone resistance genes tested, only qnrS (4.9%) was detected. Most Salmonella isolates harbored invA (100%), prgH (91.8%), and sipB (91%). Positive associations between resistance and virulence genes were observed for bla(PSE-1)/orgA, cmlA/spaN, tolC, and sul1/tolC (p < 0.05). The results suggest that companion dogs and cats are potential sources of S. enterica strains that carry resistance and virulence genes and that antimicrobial use in companion animals may select for the examined Salmonella virulence factors.

Susceptibility to β-lactams and quinolones of Enterobacteriaceae isolated from urinary tract infections in outpatients

Abstract The antibiotic susceptibility profile was evaluated in 71 Enterobacteriaceae isolates obtained from outpatient urine cultures in July 2010 from two health institutions in Santa Fe, Argentina. The highest rates of antibiotic resistance were observed for ampicillin (AMP) (69%), trimethoprim/sulfamethoxazole (TMS) (33%), and ciprofloxacin (CIP) (25%). Meanwhile, 21% of the isolates were resistant to three or more tested antibiotics families. Thirty integron-containing bacteria (42.3%) were detected, and a strong association with TMS resistance was found. Third generation cephalosporin resistance was detected in only one Escherichia coli isolate, and it was characterized as a blaCMY-2 carrier. No plasmid-mediated quinolone resistance (PMQR) was found. Resistance to fluoroquinolone in the isolates was due to alterations in QRDR regions. Two mutations in GyrA (S83L, D87N) and one in ParC (S80I) were observed in all CIP-resistant E. coli. It was determined to be the main phylogenetic groups in E. coli isolates. Minimum Inhibitory Concentration (MIC) values against nalidixic acid (NAL), levofloxacin (LEV), and CIP were determined for 63 uropathogenic E. coli isolates as MIC50 of 4 μg/mL, 0.03125 μg/mL, and 0.03125 μg/mL, respectively, while the MIC90 values of the antibiotics were determined as 1024 μg/mL, 64 μg/mL, and 16 μg/mL, respectively. An association between the phylogenetic groups, A and B1 with fluoroquinolone resistance was observed. These results point to the importance of awareness of the potential risk associated with empirical treatment with both the families of antibiotics.

Antibiotic resistance and integrons in Shiga toxin-producing Escherichia coli (STEC)

Shiga toxin-producing Escherichia coli (STEC) cause hemorrhagic colitis (HC) and hemolytic-uremic syndrome in humans (HUS). Cattle are the main reservoir of STEC and transmission to humans occurs through contaminated food and water. Antibiotics are used in pig production systems to combat disease and improve productivity and play a key role in the dissemination of antibiotic resistance genes to the bacteria. Integrons have been identified in resistant bacteria allowing for the acquisition and dissemination of antibiotic resistance genes. STEC strains isolated from humans and animals have developed antibiotic resistance. In our laboratory, 21 non-157 STEC strains isolated from pigs were analyzed to detect class 1 and 2 integrons by PCR. Eight carried integrons, 7 of them harbored intl2. In another study 545 STEC strains were also analyzed for the presence of intl1 and intl2. Strains carrying intl1 belonged to isolates from environment (n = 1), chicken hamburger (n = 2), dairy calves (n = 4) and pigs (n = 8). Two strains isolated from pigs harbored intl2 and only one intl1/intl2, highlighting the presence of intl2 in pigs. The selection for multiresistant strains may contribute to the emergence of antibiotic resistant pathogens and facilitate the spreading of the mobile resistance elements to other bacteria.

High prevalence of class 1 integrons in clinical isolates of methicillin‑resistant Staphylococcus aureus from India.

Introduction: Class1 integrons are one of the prevalent mechanisms of antibiotic resistance gene transfer in Gram‑negative organisms, but their prevalence and role in the spread of antibiotic resistance genes in methicillin‑resistant Staphylococcus aureus (MRSA) is unexplored. The purpose of this study was to investigate the prevalence of class 1 integrons in clinical isolates of MRSA. Materials and Methods: Total 143 MRSA isolates obtained from two different cities in India (Pune and Mumbai) were characterized by biochemical tests, and the antibiotic sensitivity was performed using the Clinical and Laboratory Standards Institute (CLSI) guidelines. The presence of class 1 integrons, sul1/qacEΔ1 region of class 1 integron and mecA gene among these isolates was determined by polymerase chain reaction (PCR). Results: All 143 isolates were mecA positive and coagulase‑positive. Overall, 71% of the MRSA isolates carried class 1 integrons; 58% (45/77) of the isolates obtained from Mumbai and 85% (56/66) of the isolates from Pune carried class 1 integrons. In all, 39% of these isolates carried sul1/qacEΔ1 region, thus confirming the association of class 1 integrons with antibiotic resistance genes. Along with ‑lactam antibiotics the MRSA isolates were resistant to several other antibiotics, with resistance to erythromycin, ciprofloxacin and trimethoprim‑sulfamethoxazole being observed in 75%, 66% and 60% of the isolates, respectively. Conclusion: To the best of our knowledge, this is the first report of class 1 integrons in MRSA isolates from India. The study provides insights into the prevalence of a novel mechanism adapted by MRSA for the propagation of antibiotic resistance genes.

Carriage rates of class 1, class 2 integrons as well as ISCR1 in Shigella isolates and their relationship with drug resistance / 天津医药

Objective To explore carrying rates of class 1, class 2 integrons as well as ISCR1 in Shigella isolates and their connection with drug resistance. Methods Antibiotic sensitivities were detected by K-B disk diffusion in 159 clinical isolates. Total bacteria DNA was prepared through boiling the isolates and the DNA was then used as template for PCR am?plification. PCR, ZSCR1 and sequencing analyse of integrons were applied to all of them. Results were compared by Blast and GenBank. Results Antibiotic sensitivity results showed that in the S. flexneri strains the incidence of resistance to tet?racycline and streptomycin were 88.68%and 81.13%in the S. flexneri strains while the incidence of resistant to chloram?phenicol and trimethoprim-sulfamethoxazol were both 56.60%, and the incidence of multidrug drug resistance was 77.36%. In the sonnei strains, the incidence of resistance to ampicillin, trimethoprim-sulfamethoxazo were 97.17% and 95.28%, 83.96%and 76.42%respectively, and the incidence of multidrug resistance was 98.11%. Among all isolates, 118 were class 1 integron positive , 70 were class 2 integron positive and 89 were double positives. For those 118 isolates that are positive of class 1 integron, 23 were typical while 95 weres atypical. The gene cassettes of typical class 1 integrons contains aadA2, aa?dA1, dfrⅠ, blaoxa-10 and blaoxa-1. IntI1, aadA, blaoxa-1 and IS1 were included in the gene cassetes of the atypical class 1 integrons. Class 2 integrons positive isolates carried gene cassttes which include dfrA1, satl and aadA1. No ISCR1 was found in any isolate. Integron carriage strains were closely associated with higher rate of multiple antibiobic resistance com?pared with the organisms without integrons (90.65%,50%, P<0.05). Conlusion Class 1 and class 2 integrons were widely existence in Shigella isolates and were related to the multidrug resistance.

Loop-mediated isothermal amplification assays for screening of bacterial integrons

BACKGROUND: The occurrence and prevalence of integrons in clinical microorganisms and their role played in antimicrobial resistance have been well studied recently. As screening and detection of integrons are concerned, current diagnostic methodologies are restricted by significant drawbacks and novel methods are required for integrons detection. RESULTS: In this study, three loop-mediated isothermal amplification (LAMP) assays targeting on class 1, 2 and 3 integrons were implemented and evaluated. Optimization of these detection assays were performed, including studing on the reaction temperature, volume, time, sensitivity and specificity (both primers and targets). Application of the established LAMP assays were further verified on a total of 1082 isolates (previously identified to be 397 integron-positive and 685 integron-negative strains). According to the results, the indispensability of each primer had been confirmed and the optimal reaction temperature, volume and time were found to be 65°C, 45 min and 25 µL, respectively. As application was concerned, 361, 28 and 8 isolates carrying intI1, intI2 and intI3 yielded positive amplicons, respectively. Other 685 integron-negative bacteria were negative for the integron-screening LAMP assays, totaling the detection rate and specificity to be 100%. CONCLUSIONS: The intI1-, intI2- and intI3-LAMP assays established in this study were demonstrated to be the valid and rapid detection methodologies for the screening of bacterial integrons.

Detection of drug resistance genes in integron of Shigella sonnei from six cities of Jiangsu Province in 2011 / 中华传染病杂志

Objective To investigate the resistance status of different integrons of Shigella sonnei (S.sonnei) and to analyze the distribution of resistant genes in integrons in Jiangsu Province.Methods A total of 32 strains of S.sonnei isolated from six cities of Jiangsu Province in 2011 were collected.The antibiotic susceptibility was tested by disk diffusion method.The molecular homology was analyzed by pulsed field gel electrophoresis (PFGE).The detection and classification of integrons were achieved by analyzing the positive polymerase chain reaction (PCR) products using restriction fragment length polymorphism (RFLP).RFLP and DNA sequencing were used to analyze the resistance genes in integrons.Results Multi-drug resistance (MDR) was detected in 28 (87.5％) S.sonnei strains.The resistant rates to ampicillin,nalidixic acid and tetracycline were highest (87.5％,respectively).However,it was sensitive to norfloxacin.PFGE analysis showed that there were 3 kinds of homologous clones involving 31 strains of the 32 S.sonnei strains.Among them,2,5 and 24 strains had the same clones,respectively.Accordingly,they spread within one,two and five different cities.The detection rates of class 1,class 2 and the atypical class 1 integrons in S.sonnei were 62.5％ (20/32),81.3％ (26/32) and 21.9％ (7/32),respectively,and no class 3 integron was detected.Sequence analysis of class 1 integron variable area revealed that it contained multiple resistant genes (aacA4-cmlA1 and dfrA1-aadA 1) ; dfrA1-sat 1-aadA 1 from class 2 integron and blara-30-aadA 1 from atypical class 1 integron were also identified.Conclusions In 2011,homologous S.sonnei strains spread among different cities in Jiangsu Province.MDR strains are prevalent and integrons are widespread which mediated the emergence of MDR strains.

Multidrug-resistant Acinetobacter baumannii infection in intensive care unit patients in a hospital with building construction: is there an association? / 대한마취과학회지

BACKGROUND: Acinetobacter baumannii (A. baumannii) has emerged globally as a significant pathogen in hospitals. It is also present in soil and water. In a previous study, we discovered that the A. baumannii class 2 integron occurred most frequently. Here, we determined whether the A. baumannii class 2 integron is in the soil around our hospital, and if the soil is the cause for increasing numbers of A. baumannii infections in our intensive care unit (ICU) patients. METHODS: This cross-sectional prospective study was conducted in two ICUs at Loghman-Hakim Hospital, Tehran, Iran, from November 2012 to March 2013. Patient, soil, and hospital environment samples were collected. All isolates were identified using standard bacteriologic and biochemical methods. The phenotypes and genotypes were characterized. The standard disc diffusion method was utilized to test antimicrobial susceptibility. Integron identification was performed by multiplex polymerase chain reaction. RESULTS: A total of 42 A. baumannii clinical strains were isolated, all from patient samples; 65% of the isolated species were classified as class 2 integrons. The strains were 100% resistant to piperacillin, piperacillin-tazobactam, ceftazidime, ceftriaxone, cotrimoxazole, cefepime, ceropenem, and cefotaxime. However, all of the strains were sensitive to polymyxin B. A. baumannii was detected around the lip of one patient. CONCLUSIONS: Further research is necessary to establish a relationship between A. baumannii and soil, (especially in regards to its bioremediation), as well as to determine its importance in nosocomial infections and outbreaks in the ICU.