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Vibrio spp., known as significant marine pathogens, have become more prevalent due to global warming. Antibiotics released into the environment drive Vibrio resistance. The increasing consumption of seafood leads to more interactions between Vibrio and humans. Despite this concerning trend, there remains a lack of large-scale surveillance for Vibrio contamination across various types of food. This study isolated 4027 Vibrio strains, primarily comprising V. parahaemolyticus and V. alginolyticus, in 3581 fresh shrimp and meat products from 2013 to 2022. The Vibrio strains showed increased resistance to important antibiotics, especially ß-lactams used to treat foodborne bacterial infections. Whole genome sequencing of 591 randomly chosen strains showed a strong correlation between antibiotic resistance and genotypes in Vibrio. Notably, various ESBL genes have evolved over the past 8 years, with blaVEBs being the most dominant. Additionally, carbapenemase genes, such as blaNDM-1, have become increasingly prevalent in recent years. Various mobile genetic elements, including IncQ and IncA/C plasmids, recoverable in Vibrio, facilitate the transmission of crucial ß-lactamase genes. These data provide insights into the evolutionary traits of antimicrobial resistance in foodborne Vibrio strains over a decade. Policymakers should consider these findings when devising appropriate strategies to combat bacterial antimicrobial resistance and safeguard human health.
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ß-lactam resistance is a significant global public health issue. Outbreaks of bacteria resistant to extended-spectrum ß-lactams and carbapenems are serious health concerns that not only complicate medical care but also impact patient outcomes. The primary objective of this work was to express and purify two soluble recombinant representative serine ßlactamases using Escherichia coli strain as an expression host and pET101/D as a cloning vector. Furthermore, a second objective was to evaluate the potential, innovative, and safe use of galloylquinic acid (GQA) from Copaifera lucens as a potential ß-lactamase inhibitor.In the present study, blaCTX-M-15 and blaKPC-2 represented genes encoding for serine ß-lactamases that were cloned from parent isolates of E. coli and K. pneumoniae, respectively, and expression as well as purification were performed. Moreover, susceptibility results demonstrated that recombinant cells became resistant to all test carbapenems (MICs; 64-128 µg/mL) and cephalosporins (MICs; 128-512 µg/mL). The MICs of the tested ß-lactam antibiotics were determined in combination with 4 µg/mL of GQA, clavulanic acid, or tazobactam against E. coli strains expressing CTX-M-15 or KPC-2-ß-lactamases. Interestingly, the combination with GQA resulted in an important reduction in the MIC values by 64-512-fold to the susceptible range with comparable results for other reference inhibitors. Additionally, the half-maximal inhibitory concentration of GQA was determined using nitrocefin as a ß-lactamase substrate. Data showed that the test agent was similar to tazobactam as an efficient inhibitors of the test enzymes, recording smaller IC50 values (CTX-M-15; 17.51 for tazobactam, 28.16 µg/mL for GQA however, KPC-2; 20.91 for tazobactam, 24.76 µg/mL for GQA) compared to clavulanic acid. Our work introduces GQA as a novel non-ß-lactam inhibitor, which interacts with the crucial residues involved in ß-lactam recognition and hydrolysis by non-covalent interactions, complementing the enzyme's active site. GQA markedly enhanced the potency of ß-lactams against carbapenemase and extended-spectrum ß-lactamase-producing strains, reducing the MICs of ß-lactams to the susceptible range. The ß-lactamase inhibitory activity of GQA makes it a promising lead molecule for the development of more potent ß-lactamase inhibitors.
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Escherichia coli , Pruebas de Sensibilidad Microbiana , Inhibidores de beta-Lactamasas , beta-Lactamasas , beta-Lactamasas/metabolismo , beta-Lactamasas/genética , Inhibidores de beta-Lactamasas/farmacología , Escherichia coli/genética , Klebsiella pneumoniae/efectos de los fármacos , Klebsiella pneumoniae/enzimología , Klebsiella pneumoniae/genética , Antibacterianos/farmacología , Proteínas Bacterianas/metabolismo , Proteínas Bacterianas/genética , Proteínas Bacterianas/antagonistas & inhibidores , Carbapenémicos/farmacologíaRESUMEN
Antibiotic resistance is one of most important health concerns nowadays, and ß-lactamases are the most important resistance determinants. These enzymes, based on their structural and functional characteristics, are grouped in four categories (A, B, C and D). We have solved the structure of PIB-1, a Pseudomonas aeruginosa chromosomally-encoded ß-lactamase, in its apo form and in complex with meropenem and zinc. These crystal structures show that it belongs to the Class C ß-lactamase group, although it shows notable differences, especially in the Ω- and P2-loops, which are important for the enzymatic activity. Functional analysis showed that PIB-1 is able to degrade carbapenems but not cephalosporins, the typical substrate of Class C ß-lactamases, and that its catalytic activity increases in the presence of metal ions, especially zinc. They do not bind to the active-site but they induce the formation of trimers that show an increased capacity for the degradation of the antibiotics, suggesting that this oligomer is more active than the other oligomeric species. While PIB-1 is structurally a Class C ß-lactamase, the low sequence conservation, substrate profile and its metal-dependence, prompts us to position this enzyme as the founder of a new group inside the Class C ß-lactamases. Consequently, its diversity might be wider than expected.
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Carbapenémicos , Pseudomonas aeruginosa , Zinc , beta-Lactamasas , Pseudomonas aeruginosa/enzimología , beta-Lactamasas/química , beta-Lactamasas/metabolismo , Carbapenémicos/farmacología , Carbapenémicos/metabolismo , Carbapenémicos/química , Zinc/metabolismo , Zinc/química , Modelos Moleculares , Dominio Catalítico , Hidrólisis , Especificidad por Sustrato , Metales/metabolismo , Metales/química , Metales/farmacología , Relación Estructura-Actividad , Meropenem/farmacología , Meropenem/química , Meropenem/metabolismo , Secuencia de Aminoácidos , Cristalografía por Rayos XRESUMEN
We describe four patients with a positive culture of AmpC ß-lactamases-producing Escherichia coli (E. coli), despite the fact that our understanding of plasmid-mediated AmpC ß-lactamases (pAmpC) is currently limited. Three out of four cases of AmpC ß-lactamases-producing Escherichia coli were isolated from a urine sample, and one was from a peritoneal fluid sample. All four isolates are resistant to cefoxitin disc and were subjected to a confirmatory AmpC phenotypic test (AmpC induction test) and monoplex polymerase chain reaction (PCR) for the determination of six pAmpC genotypes (blaDHA, blaEBC, blaMOX, blaFOX, blaACC, and blaCIT). All four E. coli isolates tested negative for the AmpC induction test, while monoplex PCR analysis was positive only for the blaDHA pAmpC genotype and negative for all five other genotypes (blaEBC, blaMOX, blaFOX, blaACC, and blaCIT). A common clinical characteristic across all patients was fever. One patient was treated for perforated sigmoid diverticulitis, while the other three patients were treated for acute pyelonephritis or urinary tract infections (UTIs). Each patient improved significantly and was successfully discharged.
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Treatment options for carbapenem-resistant gram-negative bacilli (CR-GNB), especially metallo-ß-lactamase (MBL)-producing CR-GNB, are limited. Aztreonam (ATM) in combination with avibactam (AVI) has shown potential for treating MBL-producing carbapenem-resistant Enterobacterales (CREs) and Stenotrophomonas maltophilia. However, data on ATM in combination with other ß-lactamase inhibitors (BLIs) are limited. We performed a multicenter study to evaluate the in vitro activities of ATM in combination with AVI, vaborbactam (VAB), relebactam (REL), tazobactam (TAZ) as well as with their commercially available formulations against CREs and S. maltophilia using broth microdilution. AVI restored ATM activity for MBL-producing CREs (ATM: 9.8% vs ATM-AVI: 78.0%) and S. maltophilia (ATM: 0% vs ATM-AVI: 93.3%). REL also moderately restored activity of ATM in MBL-producing CREs (ATM: 9.8% vs ATM-REL: 42.7%) and S. maltophilia (ATM: 0% vs ATM-REL: 68.9%). VAB and TAZ demonstrated very limited effect on the activity of ATM against CR-GNB evaluated. The combination of ATM with ceftazidime-AVI (CAZ-AVI) demonstrated maximum activity against CREs. Although ATM-CAZ-AVI is the most potent regimen available for CREs and S. maltophilia, ATM-IMI-REL might be a reasonable alternative.
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Bacterial resistance to ß-lactam antibiotics, particularly new generation cephalosporins, is a major public health concern. In Escherichia coli, resistance to these antibiotics is mainly mediated by extended-spectrum ß-lactamases (ESBL), which complicates a range of health-threatening infections. These infections may also be biofilm-related, making them more difficult to treat because of the higher tolerance to conventional antibiotics and the host immune response. In this study, we tested as potential new drug candidates against biofilm-forming ESBL-producing E. coli four antimicrobial peptides previously shown to have antifungal properties. The peptides proved to be active in vitro at micromolar concentrations against both sensitive and ESBL-producing E. coli strains, effectively killing planktonic cells and inhibiting biofilm formation. Quantitative fluorescence intensity analysis of three-dimensional reconstructed confocal laser scanning microscopy (CLSM) images of mature biofilm treated with the most active peptide showed significant eradication and a reduction in viable bacteria, while scanning electron microscopy (SEM) revealed gross morphological alterations in treated bacteria. The screening of the investigated peptides for antibacterial and antibiofilm activity led to the selection of a leading candidate to be further studied for developing new antimicrobial drugs as an alternative treatment against microbial infections, primarily associated with biofilms.
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Leminorella grimontii strain LG-KP-E1-2-T0 was isolated from Zophobas morio larvae. It showed a susceptibility phenotype compatible with the expression of an inducible extended-spectrum ß-lactamase. The presence of a chromosomal bla gene encoding for the class A GRI-1 ß-lactamase was revealed by whole-genome sequencing. GRI-1 shared the highest amino acid identity with RIC-1 and OXY-type ß-lactamases (76-80%). Analysis of six further publicly-available L. grimontii draft genomes deposited in NCBI revealed that blaGRI-1 was always present. Core-genome analysis indicated that LG-KP-E1-2-T0 was unique from other strains. We provided the first complete genome of L. grimontii and new insights on its chromosomal ß-lactamases.
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Genoma Bacteriano , beta-Lactamasas , beta-Lactamasas/genética , Animales , Cromosomas Bacterianos/genética , Secuenciación Completa del Genoma , Antibacterianos/farmacología , Pruebas de Sensibilidad Microbiana , Genómica , Larva/microbiología , Larva/genética , Mariposas Nocturnas/microbiologíaRESUMEN
In the present study, a total of 720 samples were collected from retail raw meat from 13 upazilas in Sylhet District, Bangladesh, of which 225 samples were from cattle meat, 210 samples were from goat meat, and 285 samples were from chicken meat. Salmonella enterica serovars Typhimurium and Enteritidis were screened for extended-spectrum ß-lactamase (ESBL) genes using multiplex PCR. Among the 720 samples, Salmonella spp. was detected in 28.06% (202 out of 720) of the samples, with S. Enteritidis and S. Typhimurium were identified in 11.53% (83 out of 720) and 12.22% (88 out of 720) of the samples, respectively. It was found that all Salmonella enterica serovars isolated from cattle meat displayed multidrug resistance (MDR) based on antimicrobial susceptibility testing. Notably, a significant proportion of S. Enteritidis isolates and all S. Typhimurium isolates from goat meat demonstrated complete resistance to multiple drugs (ampicillin, cefuroxime, and ceftazidime). Regarding chicken meat, out of 89 isolates encompassing both S. Typhimurium and S. Enteritidis, 57 isolates (64.04%) exhibited MDR. Additionally, blaCTX-M-1 exhibited the highest occurrence at 15.69% for S. Typhimurium and 7.89% for S. Enteritidis in chicken meat. Moreover, blaCTX-M-9 was only detected at 3.92% for S. Enteritidis in chicken meat. Furthermore, blaOXA had the highest prevalence rate of 19.04% for S. Enteritidis and 25.80% for S. Typhimurium in cattle meat, followed by chicken meat. These findings highlight the urgency for monitoring ESBL-producing Salmonella in retail raw meat and the need for strict measure to manage antibiotic use to prevent the spread of multidrug-resistant and ESBL-producing Salmonella strains, thereby protecting humans and reducing public health risks.
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The synergetic strategy has created tremendous advantages in drug-resistance bacterial infection treatment, whereas challenges related to novel compound discovery and identifying drug-binding targets still remain. The mechanisms of antimicrobial resistance involving ß-lactamase catalysis and the degradation of ß-lactam antibiotics are being revealed, with relevant therapies promising to improve the efficacy of existing major classes of antibiotics in the foreseeable future. In this study, it is demonstrated that nordalbergin, a coumarin isolated from the wood bark of Dalbergia sissoo, efficiently potentiated the activities of ß-lactam antibiotics against methicillin-resistant Staphylococcus aureus (MRSA) by suppressing ß-lactamase performance and improving the bacterial biofilm susceptibility to antibiotics. Nordalbergin was found to destabilize the cell membrane and promote its permeabilization. Moreover, nordalbergin efficiently improved the therapeutic efficacy of amoxicillin against MRSA pneumonia in mice, as supported by the lower bacterial load, attenuated pathological damage, and decreased inflammation level. These results demonstrate that nordalbergin might be a promising synergist of amoxicillin against MRSA infections. This study provided a new approach for developing potentiators for ß-lactam antibiotics against MRSA infections.
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Antibacterianos , Sinergismo Farmacológico , Staphylococcus aureus Resistente a Meticilina , Infecciones Estafilocócicas , beta-Lactamas , Staphylococcus aureus Resistente a Meticilina/efectos de los fármacos , Animales , Antibacterianos/farmacología , Antibacterianos/química , Ratones , Infecciones Estafilocócicas/tratamiento farmacológico , Infecciones Estafilocócicas/microbiología , beta-Lactamas/farmacología , Pruebas de Sensibilidad Microbiana , Biopelículas/efectos de los fármacos , Cumarinas/farmacología , Cumarinas/química , beta-Lactamasas/metabolismo , Amoxicilina/farmacología , Antibióticos BetalactámicosRESUMEN
INTRODUCTION: Our goal was to investigate the antimicrobial resistance due to beta-lactamase genes and virulent determinants (biofilm-forming ability) expressed by Acinetobacter collected from health settings in Pakistan. A cross-sectional study was conducted for the molecular characterization of carbapenemases and biofilm-producing strains of Acinetobacter spp. METHODOLOGY: Two twenty-three imipenem-resistant Acinetobacter isolates were analyzed from 2020 to 2023.The combination disk test and modified hodge test were performed. Biofilm forming ability was determined by polystyrene tube assay. Multiplex polymerase chain reaction (PCR) for virulent and biofilm-forming genes, and 16S rRNA sequencing were performed. RESULTS: 118 (52.9%) carbapenem-resistant Acinetobacter (CR-AB) were isolated from wounds and pus, 121 (54.2%) from males, and 92 (41.2%) from 26-50-years-olds. More than 80% of strains produced ß-lactamases and carbapenemases. Based on the PCR amplification of the ITS gene, 174 (78.0%) CR-AB strains were identified from CR-Acinetobacter non-baumannii (ANB). Most CR-AB were strong and moderate biofilm producers. Genetic analysis revealed the blaOXA-23, blaTEM, blaCTX-M blaNDM-1 and blaVIM were prevalent in CR-AB with frequencies 91 (94.8%), 68 (70.8%), 19 (19.7%), 53 (55.2%), 2 (2.0%) respectively. Among virulence genes, OmpA was dominant in CR-AB isolates from wound (83, 86.4%), csuE 63 (80.7%) from non-wound specimens and significantly correlated with blaNDM and blaOXA genes. Phylogenetic analysis revealed three different clades for strains based on specimens. CONCLUSIONS: CR-AB was highly prevalent in Pakistan and associated with wound infections. The genes, blaOXA-23, blaTEM, blaCTX-M, and blaNDM-1 were detected in CR-AB. Most CR-AB were strong biofilm producers with virulent genes OmpA and csuE.
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Infecciones por Acinetobacter , Acinetobacter baumannii , Antibacterianos , Biopelículas , Carbapenémicos , beta-Lactamasas , Biopelículas/crecimiento & desarrollo , beta-Lactamasas/genética , Humanos , Pakistán , Acinetobacter baumannii/genética , Acinetobacter baumannii/efectos de los fármacos , Masculino , Estudios Transversales , Adulto , Persona de Mediana Edad , Femenino , Infecciones por Acinetobacter/microbiología , Antibacterianos/farmacología , Carbapenémicos/farmacología , Pruebas de Sensibilidad Microbiana , Adulto Joven , Proteínas Bacterianas/genética , AdolescenteRESUMEN
BACKGROUND: Currently, the Enterobacteriaceae species are responsible for a variety of serious infections and are already considered a global public health problem, especially in underdeveloped countries, where surveillance and monitoring programs are still scarce and limited. Analyses were performed on the complete genome of an extensively antibiotic-resistant strain of Enterobater hormaechei, which was isolated from a patient with non-Hodgkin's lymphoma, who had been admitted to a hospital in the city of Manaus, Brazil. METHODS: Phenotypical identification and susceptibility tests were performed in automated equipment. Total DNA extraction was performed using the PureLink genomic DNA mini-Kit. The genomic DNA library was prepared with Illumina Microbial Amplicon Prep and sequenced in the MiSeq Illumina Platform. The assembly of the whole-genome and individual analyses of specific resistance genes extracted were carried out using online tools and the Geneious Prime software. RESULTS: The analyses identified an extensively resistant ST90 clone of E. hormaechei carrying different genes, including blaCTX-M-15, blaGES-2, blaTEM-1A, blaACT-15, blaOXA-1 and blaNDM-1, [aac(3)-IIa, aac(6')-Ian, ant(2â³)-Ia], [aac(6')-Ib-cr, (qnrB1)], dfrA25, sul1 and sul2, catB3, fosA, and qnrB, in addition to resistance to chlorhexidine, which is widely used in patient antisepsis. CONCLUSIONS: These findings highlight the need for actions to control and monitor these pathogens in the hospital environment.
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Farmacorresistencia Bacteriana Múltiple , Enterobacter , Genoma Bacteriano , Linfoma no Hodgkin , Secuenciación Completa del Genoma , Humanos , Enterobacter/genética , Enterobacter/efectos de los fármacos , Enterobacter/aislamiento & purificación , Linfoma no Hodgkin/genética , Linfoma no Hodgkin/microbiología , Linfoma no Hodgkin/tratamiento farmacológico , Farmacorresistencia Bacteriana Múltiple/genética , Secuenciación Completa del Genoma/métodos , Antibacterianos/farmacología , Antibacterianos/uso terapéutico , Infecciones por Enterobacteriaceae/microbiología , Infecciones por Enterobacteriaceae/tratamiento farmacológico , Infecciones por Enterobacteriaceae/genética , Pruebas de Sensibilidad Microbiana , BrasilRESUMEN
New ß-lactam-ß-lactamase inhibitor combinations represent last-resort antibiotics to treat infections caused by multidrug-resistant Pseudomonas aeruginosa. Carbapenemase gene acquisition can limit their spectrum of activity, and reports of resistance toward these new molecules are increasing. In this multi-center study, we evaluated the prevalence of resistance to ceftazidime-avibactam (CZA) and comparators among P. aeruginosa clinical isolates from bloodstream infections, hospital-acquired or ventilator-associated pneumonia, and urinary tract infections, circulating in Southern Italy. We also investigated the clonality and content of relevant ß-lactam resistance mechanisms of CZA-resistant (CZAR) isolates. A total of 120 P. aeruginosa isolates were collected. CZA was among the most active ß-lactams, retaining susceptibility in the 81.7% of cases, preceded by cefiderocol (95.8%) and followed by ceftolozane-tazobactam (79.2%), meropenem-vaborbactam (76.1%), imipenem-relebactam (75%), and aztreonam (69.6%). Among non-ß-lactams, colistin and amikacin were active against 100% and 85.8% of isolates respectively. In CZAR strains subjected to whole-genome sequencing (n = 18), resistance was mainly due to the expression of metallo-ß-lactamases (66.6% VIM-type and 5.5% FIM-1), followed by PER-1 (16.6%) and GES-1 (5.5%) extended-spectrum ß-lactamases, mostly carried by international high-risk clones (ST111 and ST235). Of note, two strains producing the PER-1 enzyme were resistant to all ß-lactams, including cefiderocol. In conclusion, the CZA resistance rate among P. aeruginosa clinical isolates in Southern Italy remained low. CZAR isolates were mostly metallo-ß-lactamases producers and belonging to ST111 and ST253 epidemic clones. It is important to implement robust surveillance systems to monitor emergence of new resistance mechanisms and to limit the spread of P. aeruginosa high-risk clones. IMPORTANCE: Multidrug-resistant Pseudomonas aeruginosa infections are a growing threat due to the limited therapeutic options available. Ceftazidime-avibactam (CZA) is among the last-resort antibiotics for the treatment of difficult-to-treat P. aeruginosa infections, although resistance due to the acquisition of transferable ß-lactamase genes is increasing. With this work, we report that CZA represents a highly active antipseudomonal ß-lactam compound (after cefiderocol), and that metallo-ß-lactamases (VIM-type) and extended-spectrum ß-lactamases (GES and PER-type) production is the major factor underlying CZA resistance in isolates from Southern Italian hospitals. In addition, we reported that such resistance mechanisms were mainly carried by the international high-risk clones ST111 and ST235.
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Antibacterianos , Compuestos de Azabiciclo , Ceftazidima , Combinación de Medicamentos , Farmacorresistencia Bacteriana Múltiple , Pruebas de Sensibilidad Microbiana , Infecciones por Pseudomonas , Pseudomonas aeruginosa , Compuestos de Azabiciclo/farmacología , Ceftazidima/farmacología , Humanos , Pseudomonas aeruginosa/efectos de los fármacos , Pseudomonas aeruginosa/genética , Pseudomonas aeruginosa/aislamiento & purificación , Antibacterianos/farmacología , Infecciones por Pseudomonas/microbiología , Infecciones por Pseudomonas/tratamiento farmacológico , Infecciones por Pseudomonas/epidemiología , Farmacorresistencia Bacteriana Múltiple/genética , Italia/epidemiología , Inhibidores de beta-Lactamasas/farmacología , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Masculino , Femenino , beta-Lactamasas/genética , beta-Lactamasas/metabolismo , Persona de Mediana Edad , beta-Lactamas/farmacología , Anciano , AdultoRESUMEN
AIMS: To investigate the genetic profile and characterize antimicrobial resistance, including the main ß-lactam antibiotic resistance genes, in Acinetobacterbaumannii isolates from a tertiary hospital in Recife-PE, Brazil, in the post-COVID-19 pandemic period. METHODS AND RESULTS: Acinetobacter baumannii isolates were collected between 2023 and 2024 from diverse clinical samples. Antimicrobial resistance testing followed standardized protocols, with ß-lactamase-encoding genes detected via PCR and sequencing. Investigation into ISAba1 upstream of blaOXA-carbapenemase and blaADC genes was also conducted. Genetic diversity was assessed through ERIC-PCR. Among the 78 A. baumannii, widespread resistance to multiple antimicrobials was evident. Various acquired ß-lactamase-encoding genes (blaOXA-23,-24,-58,-143, blaVIM, and blaNDM) were detected. Furthermore, this is the first report of blaVIM-2 in A. baumannii isolates harboring either the blaOXA-23-like or the blaOXA-143 gene in Brazil. Molecular typing revealed a high genetic heterogeneity among the isolates, and multi-clonal dissemination. CONCLUSION: The accumulation of genetic resistance determinants underscores the necessity for stringent infection control measures and robust antimicrobial stewardship programs to curb multidrug-resistant strains.
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Infecciones por Acinetobacter , Acinetobacter baumannii , Antibacterianos , COVID-19 , Pruebas de Sensibilidad Microbiana , SARS-CoV-2 , Centros de Atención Terciaria , beta-Lactamasas , Acinetobacter baumannii/genética , Acinetobacter baumannii/efectos de los fármacos , Acinetobacter baumannii/aislamiento & purificación , Brasil , Humanos , Infecciones por Acinetobacter/microbiología , Infecciones por Acinetobacter/tratamiento farmacológico , Infecciones por Acinetobacter/epidemiología , beta-Lactamasas/genética , Antibacterianos/farmacología , Antibacterianos/uso terapéutico , SARS-CoV-2/genética , Farmacorresistencia Bacteriana Múltiple/genética , Proteínas Bacterianas/genética , Masculino , Adulto , Femenino , Persona de Mediana Edad , Farmacorresistencia Bacteriana/genéticaRESUMEN
BACKGROUND AND OBJECTIVE(S): CRISPR-Cas is a prokaryotic adaptive immune system that protects bacteria and archaea against mobile genetic elements (MGEs) such as bacteriophages plasmids, and transposons. In this study, we aimed to assess the prevalence of the CRISPR-Cas systems and their association with antibiotic resistance in one of the most challenging bacterial pathogens, Klebsiella pneumoniae. MATERIALS AND METHODS: A total of 105 K. pneumoniae isolates were collected from various clinical infections. Extended-spectrum ß-lactamases (ESBLs) phenotypically were detected and the presence of ESBL, aminoglycoside-modifying enzymes (AME), and CRISPR-Cas system subtype genes were identified using PCR. Moreover, the diversity of the isolates was determined by enterobacterial repetitive intergenic consensus (ERIC)-PCR. RESULTS: Phenotypically, 41.9% (44/105) of the isolates were found to be ESBL producers. A significant inverse correlation existed between the subtype I-E CRISPR-Cas system's presence and ESBL production in K. pneumoniae isolates. Additionally, the frequency of the ESBL genes blaCTX-M1 (3%), blaCTX-M9 (12.1%), blaSHV (51.5%), and blaTEM (33.3%), as well as some AME genes such as aac(3)-Iva (21.2%) and ant(2'')-Ia (3%) was significantly lower in the isolates with the subtype I-E CRISPR-Cas system in comparison to CRISPR-negative isolates. There was a significant inverse correlation between the presence of ESBL and some AME genes with subtype I-E CRISPR-Cas system. CONCLUSION: The presence of the subtype I-E CRISPR-Cas system was correlated with the antibiotic-resistant gene (ARGs). The isolates with subtype I-E CRISPR-Cas system had a lower frequency of ESBL genes and some AME genes than CRISPR-negative isolates.
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Antibacterianos , Sistemas CRISPR-Cas , Infecciones por Klebsiella , Klebsiella pneumoniae , beta-Lactamasas , Klebsiella pneumoniae/genética , Klebsiella pneumoniae/efectos de los fármacos , Humanos , beta-Lactamasas/genética , Infecciones por Klebsiella/microbiología , Infecciones por Klebsiella/epidemiología , Antibacterianos/farmacología , Pruebas de Sensibilidad Microbiana , Farmacorresistencia Bacteriana/genética , Prevalencia , Masculino , Femenino , Persona de Mediana EdadRESUMEN
Background and Aim: Established antimicrobial resistance (AMR) surveillance in companion animals is lacking, particularly in low-middle-income countries. The aim of this study was to analyze AMR and its risk factors in Escherichia coli isolated from dogs at two veterinary centers in Lima (Peru). Materials and Methods: Ninety dogs were included in the study. Antimicrobial susceptibility was established by disk diffusion, whereas microdilution was used to determine colistin susceptibility. Mechanisms related to extended-spectrum ß-lactamases (ESBL) and colistin resistance were determined by polymerase chain reaction. Clonal relationships of colistin-resistant isolates were assessed by XbaI-pulsed-field gel electrophoresis. Results: Thirty-five E. coli strains were isolated. High levels of resistance to ampicillin (57.1%), nalidixic acid (54.3%), tetracycline (48.6%), and azithromycin (25.7%) were detected. Cephalosporin resistance levels were ≥20% and those for colistin were 14.3%. Twelve (34.2%) isolates were ESBL producers; of these, six blaCTX-M-55 (50.0%), 2 (16.6%) blaCTX-M-15, and 2 (16.6%) blaCTX-M-8-like genes were found. The five colistin-resistant isolates were clonally unrelated, with four of them presenting amino acid codon substitutions in the mgrB gene (V8A) or mutations in the mgrB promoter (a12g, g98t, and c89t). Furthermore, dog age, <6 years (p = 0.027) and raw diet (p = 0.054) were associated with resistance to a greater number of antibiotic families. Conclusion: Despite small number of samples included, the study found that dogs studied were carriers of multidrug-resistant E. coli, including last-resort antimicrobials, representing a public health problem due to close contact between dogs and humans. This issue suggests the need for larger studies addressed to design strategies to prevent the spread of resistant micro-organisms in small animal clinics and domestic settings.
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In this study, we proposed a novel method utilizing polyethyleneimine (PEI)-modified halloysite nanotubes (HNTs)-based hybrid silica monolithic spin tip to analyze hydrophilic ß-lactam antibiotics and ß-lactamases inhibitors in whole blood samples for the first time. HNTs were incorporated directly into the hybrid silica monolith via a sol-gel method, which improved the hydrophilicity of the matrix. The as-prepared monolith was further modified with PEI by glutaraldehyde coupling reaction. It was found that the PEI-modified HNTs-based hybrid silica monolith enabled a large adsorption capacity of cefoperazone at 35.7 mg g-1. The monolithic spin tip-based purification method greatly reduced the matrix effect of whole blood samples and had a detection limit as low as 0.1 - 0.2 ng mL-1. In addition, the spiked recoveries of sulbactam, cefuroxime, and cefoperazone in blank whole blood were in the range of 89.3-105.4 % for intra-day and 90.6-103.5 % for inter-day, with low relative standard deviations of 1.3-7.2 % and 4.9-10.5 %, respectively. This study introduces a new strategy for preparing nanoparticles incorporated in a hybrid silica monolith with a high adsorption capacity. Moreover, it offers a valuable tool to monitor sulbactam, cefoperazone, and cefuroxime in whole blood from pregnant women with the final aim of guiding their administration.
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Cefoperazona , Cefuroxima , Interacciones Hidrofóbicas e Hidrofílicas , Límite de Detección , Nanotubos , Dióxido de Silicio , Extracción en Fase Sólida , Sulbactam , Cefoperazona/sangre , Cefoperazona/química , Humanos , Sulbactam/sangre , Sulbactam/química , Extracción en Fase Sólida/métodos , Dióxido de Silicio/química , Nanotubos/química , Cefuroxima/sangre , Cefuroxima/química , Arcilla/química , Adsorción , Antibacterianos/sangre , Antibacterianos/química , Polietileneimina/química , Cromatografía Líquida de Alta Presión/métodos , Reproducibilidad de los ResultadosRESUMEN
The spread of superbugs in dairy products can jeopardize global public health. To date, information on the incidence rates of virulent and ß-lactams-resistant (BLR) Salmonella in cheeses from rural areas of Egypt has been lacking. Biochemical, serological, antibiotic susceptibility, and multiplex PCR (M-PCR) tests were performed to identify and characterize Salmonella isolates. In this study, 44 (15.71%) Salmonella isolates of eight different serotypes were recovered from 280 samples of Rumi and pasteurized Kariesh cheeses across the Nile Delta region of Egypt. The most predominant serotypes were S. Typhimurium, S. Enteritidis, and S. Infantis. The virulence genes (invA, stn, and hilA) were identified in all isolates. However, spvC was only detected in S. Typhimurium. The highest resistance was developed against Erythromycin and Clindamycin (90.91%), followed by Ceftazidime and Cephalothin (84.09%). Meropenem and colistin were the most effective antibiotics. A high proportion (79.55%) of multi-drug resistance (MDR) isolates carried narrow spectrum (NS), extended-spectrum (ES), and AmpC-BLR genes. The blaOXA-1, blaOXA-2, blaTEM-1, blaCTX-M, blaCMY-1, and blaCMY-2 BLR genes were positive in 37.04%, 29.63%, 25.93%, 14.81%, 37.04%, and 3.70% of isolates, respectively. In conclusion, a high prevalence of virulence and BLR genes harboring Salmonella strains in Egyptian cheeses is considered a great threat to public health.
RESUMEN
Beta-lactamase-producing Enterobacterales bacteria cause severe hard-to-treat infections. Currently, they are spreading beyond hospitals and becoming a serious global health concern. This study investigated the prevalence and molecular characterization of extended-spectrum ß-lactamase and AmpC-type ß-lactamase-producing Enterobacterales (ESBL-PE, AmpC-PE) in wastewater from livestock and poultry slaughterhouses in Ardabil, Iran. A total of 80 Enterobacterales bacteria belonging to 9 species were identified. Among the isolates, Escherichia coli (n = 21/80; 26.2%) and Citrobacter spp. (n = 18/80; 22.5%) exhibited the highest frequency. Overall, 18.7% (n = 15/80) and 2.5% (n = 2/80) of Enterobacterales were found to be ESBL and AmpC producers, respectively. The most common ESBL producer isolates were E. coli (n = 9/21; 42.8%) and Klebsiella pneumoniae (n = 6/7; 85.7%). All AmpC-PE isolates belonged to E. coli strains (n = 2/21; 9.5%). In this study, 80% of ESBL-PE and 100% of AmpC-PE isolates were recovered from poultry slaughterhouse wastewater. All ESBL-PE and AmpC-PE isolates were multidrug-resistant. In total, 93.3% of ESBL-PE isolates harbored the blaCTX-M gene, with the blaCTX-M-15 being the most common subgroup. The emergence of ESBL-PE and AmpC-PE in wastewater of food-producing animals allows for zoonotic transmission to humans through contaminated food products and contaminations of the environment.
Asunto(s)
Infecciones por Escherichia coli , Escherichia coli , Animales , Humanos , Infecciones por Escherichia coli/epidemiología , Infecciones por Escherichia coli/microbiología , Aves de Corral/microbiología , Mataderos , Ganado , Aguas Residuales , Prevalencia , Irán , Antibacterianos , beta-Lactamasas/genética , Proteínas Bacterianas/genética , BacteriasRESUMEN
ß-Lactamases (EC 3.5.2.6) confer resistance against ß-lactam group-containing antibiotics in bacteria and higher eukaryotes, including humans. Pathogenic bacterial resistance against ß-lactam antibiotics is a primary concern for potential therapeutic developments and drug targets. Here, we report putative ß-lactamase activity, sulbactam binding (a ß-lactam analogue) in the low µM affinity range, and site-specific interaction studies of a 14 kDa UV- and dark-inducible protein (abbreviated as UVI31+, a BolA homologue) from Chlamydomonas reinhartii. Intriguingly, the solution NMR structure of UVI31 + bears no resemblance to other known ß-lactamases; however, the sulbactam binding is found at two sites rich in positively charged residues, mainly at the L2 loop regions and the N-terminus. Using NMR spectroscopy, ITC and MD simulations, we map the ligand binding sites in UVI31 + providing atomic-level insights into its ß-lactamase activity. Current study is the first report on ß-lactamase activity of UVI31+, a BolA analogue, from C. reinhartii. Furthermore, our mutation studies reveal that the active site serine-55 is crucial for ß-lactamase activity.
Asunto(s)
Chlamydomonas reinhardtii , beta-Lactamasas , Chlamydomonas reinhardtii/enzimología , beta-Lactamasas/química , beta-Lactamasas/metabolismo , Sitios de Unión , Resonancia Magnética Nuclear Biomolecular/métodos , Sulbactam/química , Sulbactam/farmacología , Espectroscopía de Resonancia Magnética/métodos , Simulación de Dinámica Molecular , Secuencia de Aminoácidos , Proteínas de Plantas/química , Proteínas de Plantas/metabolismo , Unión ProteicaRESUMEN
Developing simple, inexpensive, fast, sensitive, and specific probes for antibiotic-resistant bacteria is crucial for the management of urinary tract infections (UTIs). We here propose a paper-based sensor for the rapid detection of ß-lactamase-producing bacteria in the urine samples of UTI patients. By conjugating a strongly electronegative group -N+(CH3)3 with the core structures of cephalosporin and carbapenem antibiotics, two visual probes were achieved to respectively target the extended-spectrum/AmpC ß-lactamases (ESBL/AmpC) and carbapenemase, the two most prevalent factors causing antibiotic resistance. By integrating these probes into a portable paper sensor, we confirmed 10 and 8 cases out of 30 clinical urine samples as ESBL/AmpC- and carbapenemase-positive, respectively, demonstrating 100% clinical sensitivity and specificity. This paper sensor can be easily conducted on-site, without resorting to bacterial culture, providing a solution to the challenge of rapid detection of ß-lactamase-producing bacteria, particularly in resource-limited settings.