Primer reporte de Enterobacterales dobles productores de carbapenemasas en hospitales de Paraguay. Año 2021

RESUMEN Las carbapenemasas se encuentran ampliamente distribuidas en nuestro país, tanto en bacilos gramnegativos fermentadores como no fermentadores. Durante 2021, se ha reportado incremento de cepas con estas enzimas. Con el objetivo de evaluar la doble producción de carbapenemasas en Enterobacterales y comunicar su circulación, fue puesta a punto una PCR convencional múltiple. Estudio retrospectivo en 128 aislamientos provenientes de 20 centros colaboradores de la Red Nacional de Vigilancia de la RAM (Capital, Central e interior del país), remitidos al LCSP entre febrero y setiembre de 2021, para confirmación y genotipificación de carbapenemasas. Se realizaron pruebas fenotípicas y colorimétricas con sustratos específicos, y pruebas genotípicas (PCR convencional múltiple) para la detección simultánea de varios genes de resistencia (bla NDM, bla KPC, bla OXA-48-like, bla IMP y bla VIM). De los 128 aislamientos estudiados, 107 correspondieron a Klebsiella pneumoniae, 14 a Enterobacter cloacae complex, entre otros; aislados en mayor frecuencia de muestras de orina (30%), respiratorias (30%), sangre y catéter (24%). Los genes de resistencia a los carbapenemes detectados fueron: bla NDM (77,3%), bla KPC (17,2%); siendo confirmada la doble producción de carbapenemasas en 7 aislamientos (5,5%) provenientes de 4 centros diferentes de la capital de país y uno de Central; 6 de ellas (K. pneumoniae) con bla NDM+bla KPC y 1 (E. cloacae complex) con bla NDM+bla OXA-48-like; confirmando circulación de Enterobacterales dobles productores de carbapenemasas en el país (KPC+NDM y OXA+NDM); hallazgos que obligan a proveer de capacidades de detección, de manera a que se puedan tomar medidas oportunas y eficaces de contención y control.

ABSTRACT Carbapenemases are widely distributed in our country, both in fermenting and non-fermenting gram-negative bacilli. During 2021, an increase in strains with these enzymes has been reported. In order to evaluate the double production of carbapenemases in Enterobacterales and communicate their circulation, a multiple conventional PCR was set up. Retrospective study carried out in 128 isolates from 20 collaborating centers of the National AMR Surveillance Network (Capital, Central and interior of the country), sent to the LCSP between February and September 2021, for confirmation and genotyping of carbapenemases. Phenotypic and colorimetric tests were performed with specific substrates, as well as genotypic tests (multiple conventional PCR) for the simultaneous detection of several resistance genes (blaNDM, blaKPC, blaOXA-48-like, blaIMP and blaVIM). Of the 128 isolates studied, 107 corresponded to Klebsiella pneumoniae, 14 to Enterobacter cloacae complex, among others; isolated in higher frequency from urine (30%), respiratory (30%), blood and catheter (24%) samples. The genes for resistance to carbapenems detected were: blaNDM (77.3%), blaKPC (17.2%); the double production of carbapenemases was confirmed in 7 isolates (5.5%) from 4 different centers in the capital of the country and one in Central; 6 of them (K. pneumoniae) with blaNDM + blaKPC and 1 (E. cloacae complex) with blaNDM + blaOXA-48-like; confirming circulation of double Enterobacterales producers of carbapenemases in the country (KPC + NDM and OXA + NDM); findings that require the provision of detection capabilities, so that timely and effective containment and control measures can be taken.

First report of a blaNDM-resistant gene in a Klebsiella aerogenes clinical isolate from Brazil

Abstract INTRODUCTION: Carbapenemase-resistant enterobacteria that produce the bla NDM gene are found worldwide. However, this is the first report of blaNDM in Klebsiella aerogenes in Brazil. METHODS: The identification of bacterial species was performed using anautomated system and confirmed by biochemical tests, 16S rRNA gene sequencing, and detection of resistance genes. RESULTS: The clinical isolate showed minimum inhibitory concentration resistance to meropenem and polymyxin B at 8mg/L and 4mg/L, respectively. Only the blaNDM gene was detected. CONCLUSIONS: The current report of the blaNDM gene in isolated MDR enterobacteria indicates that this gene can spread silently in a hospital setting.

First report of bla GES-1 in Proteus mirabilis clinical isolates

Abstract Proteus mirabilis is one of the main pathogens causing urinary tract infections and sepsis. To our knowledge, this is the first report of a P. mirabilis hosting bla GES. The presence of these genes was determined using PCR and sequencing. We identified the presence of bla GES-1 in all three isolates. In addition, we identified the bla KPC-2 and bla NDM-1 genes in the two strains. These data emphasize the importance of monitoring and surveillance of all enterobacteria. The circulation of P. mirabilis strains carrying bla GES-1 constitutes a new scenario of resistance in this species and should be an epidemiological alert for global health.

Will ceftazidime/avibactam plus aztreonam be effective for NDM and OXA-48-Like producing organisms: Lessons learnt from In vitro study

Introduction: Carbapenem resistance (CR) in Klebsiella pneumoniae is mainly mediated by bla NDM and bla OXA-48 carbapenemases. Newer Food and Drug Administration-approved antimicrobial ceftazidime/avibactam (C/A) has a potent activity against bla OXA-48-like producers. However, its activity is limited in organisms co-producing bla NDM and bla OXA-48-like. Addition of aztreonam (ATM) to C/A potentially expands the spectrum of coverage for carbapenemase co-producers. With this, we aimed to determine the synergistic activity of combination of C/A plus ATM against bla NDM, bla OXA-48-like and co-producers of bla NDM + bla OXA-48-like producing CR Klebsiella pneumoniae (CRKp). Materials and Methods: A total of 12 isolates of CRKp-harbouring genes encoding bla NDM and bla OXA-48-like were tested. Minimum inhibitory concentrations (MICs) were determined for several antimicrobial agents, including C/A (0.5�?g/ml) by broth microdilution method. Checkerboard assay was performed for the combination of C/A plus ATM at varying concentrations. Fold differences in the MIC of C/A with and without addition of ATM were determined to infer synergistic effects. Results: MIC of C/A and ATM ranged from 0.5 to >8 ?g/ml and 64 to 2048 ?g/ml, respectively. Two isolates were susceptible to C/A with MIC of 0.5 and 1 ?g/ml, while others were resistant with MIC of >8 ?g/ml. Synergistic effects of >8-fold MIC difference in C/A MIC were noted with addition of ATM at 4 ?g/ml. This was observed for all CRKp with profiles of bla NDM, bla OXA-48-like and co-producers of bla NDM + bla OXA-48-like genes, which was a promising effect. Notably, all five of the colistin-resistant CRKp were inhibited with >8-fold MIC difference in the combination of C/A plus ATM at 4 ?g/ml. Conclusion: With the increasing burden of CRKp, the use of C/A with ATM combination seems to be very promising, especially for bla NDM, bla OXA-48-like and co-producers of bla NDM + bla OXA-48like carbapenemases.

Emergence of bla NDM-1 associated with the aac(6')-Ib-cr, acrB, cps, and mrkD genes in a clinical isolate of multi-drug resistant Klebsiella pneumoniae from Recife-PE, Brazil

Abstract INTRODUCTION: The emergence of New Delhi metallo-β-lactamase (NDM) is concernig because it reduces the antibiotic therapy options for bacterial infections. METHODS: Resistant and virulent genes from an isolate of Klebsiella pneumoniae derived from a patient with sepsis in a hospital in Recife-PE, Brazil, were investigated using PCR and DNA sequencing. RESULTS: bla NDM-1, aac(6')-Ib-cr and acrB resistance genes, and cps and mrkD virulence genes were detected. CONCLUSIONS To our knowledge, this is the first report on bla NDM-1 in Recife-PE. This detection alerts researchers to the need to control the spread of bla NDM-1 resistance gene by this bacterium in Brazil.

NDM-1 (New Delhi metallo beta lactamase-1) producing Gram-negative bacilli: Emergence & clinical implications.

Backgound & objectives: resistance to carbapenems in Gram-negative bacteria conferred by NDM-1 is a global health problem. We investigated the occurrence of NDM-1 in clinical isolates of gram-negative bacilli in a tertiary care hospital in Kashmir valley, India. Methods: Gram-negative bacilli from different clinical isolates were included in the study. Antimicrobial susceptibility was performed by Kirby Bauer disk diffusion method and interpreted using Clinical Laboratory Standards Institute (CLSI) guidelines. Isolates resistant to carbapenems were subjected to different phenotypic test such as modified hodge test (MHT), boronic acid and oxacillin based MHT (bA-MHT and OXA-MHT), combined disk test and minimum inhibitory concentration (MIC) with imipenem and imipenem -EDTA for determination of class B metallo enzymes. Presence of blaNDM-1 gene was established by PCR and confirmed by sequencing. Results: Of the total 1625 gram-negative isolates received, 100 were resistant to imipenem. Of the 100 isolates, 55 (55%) were positive by modified Hodge test indicating carbapenemase production. Of the 100 isolates tested by MHT, BA-MHT and OXA-MHT, 29 (29%) isolates belonged to Class A and 15 (15%) to Class B, while 56 (56%) isolates were negative. Of the 15 class B metallo beta lactamase producers, nine carried the blaNDM-1 gene. NDM-1 was found among escherichia coli (2 isolates), Klebsiella pneumoniae (2 isolates), Citrobacter freundii (3 isolates), Acinetobacter spp (1 isolate), and one isolate of Pseudomonas aeruginosa. Isolates were resistant to all antibiotic tested except polymyxin B and tigecycline. Interpretation & conclusions: Our study showed the presence of clinical isolates expressing NDM-1 in Srinagar, Jammu & Kashmir, India. These isolates harbour plasmid mediated multiple drug resistant determinants and can disseminate easily across several unrelated genera. To halt their spread, early identification of these isolates is mandatory.