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Pseudomonas aeruginosa is a leading cause of hospital-acquired infections worldwide. Biofilm production, antibiotic resistance, and a wide range of virulence factors contribute to their persistence in nosocomial environments. We describe an outbreak caused by a multidrug-resistant P. aeruginosa strain in an ICU. Antibiotic susceptibility was determined and blaPER-1 and qnrVC were amplified via PCR. Clonality was determined using PFGE and biofilm formation was studied with a static model. A combination of antibiotics was assessed on both planktonic cells and biofilms. WGS was performed on five isolates. All isolates were clonally related, resistant to ceftazidime, cefepime, amikacin, and ceftolozane-tazobactam, and harbored blaPER-1; 11/19 possessed qnrVC. Meropenem and ciprofloxacin reduced the biofilm biomass; however, the response to antibiotic combinations with rifampicin was different between planktonic cells and biofilms. WGS revealed that the isolates belonged to ST309 and serotype O11. blaPER-1 and qnrVC6 were associated with a tandem of ISCR1 as part of a complex class one integron, with aac(6')-Il and ltrA as gene cassettes. The structure was associated upstream and downstream with Tn4662 and flanked by direct repeats, suggesting its horizontal mobilization capability as a composite transposon. ST309 is considered an emerging high-risk clone that should be monitored in the Americas.
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Antibiotic resistance is an alarming problem throughout the world and carbapenem-resistant Pseudomonas aeruginosa has been cataloged as critical in the World Health Organization list of microorganisms in urgent need for the development of new antimicrobials. In this work, we describe two novel resistance regions responsible for conferring a multidrug resistance phenotype to two clinical isolates of P. aeruginosa (Pa873 and Pa6415) obtained from patients hospitalized in the ICU of University Hospital of Uruguay. Bacterial identification and antibiotic susceptibility tests were performed using MALDI-TOF and the Vitek 2 system, respectively. WGS was performed for both isolates using Oxford Nanopore Technologies and Illumina and processed by means of hybrid assembly. Both isolates were resistant to ceftazidime, cefepime, piperacillin-tazobactam, aztreonam, and imipenem. Strain Pa6415 also showed resistance to ciprofloxacin. Both strains displayed MICs below the susceptibility breakpoint for CAZ-AVI plus 4 mg/L of aztreonam as well as cefiderocol. Both resistance regions are flanked by the left and right inverted repeats of ISPa40 in two small regions spanning 39.3 and 35.6 kb, for Pa6415 and Pa873, respectively. The resistance region of Pa6415 includes TnaphA6, and the new Tn7516 consists of IRi, In899, qacEΔ1-sul1-ISCR1, qnrVC6-ISCR1-blaPER-1-qacEΔ1-sul1, araJ-like, IS481-like tnpA, ISPa17, and IRR. On the other hand, the resistance region of Pa873 includes Tnaph6 and the new Tn7517 (IRi, In899, qacEΔ1-sul1, ISCR1-blaPER-1-qacEΔ1-sul1, araJ-like, IS481-like tnpA, ISPa17, and IRR). It is necessary to monitor the emergence of genetic structures that threaten to invalidate the available therapeutic resources.
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Resumen: A nivel mundial se estima que en 2018 hubo alrededor de 10 millones de nuevos casos de tuberculosis (TBC). La detección molecular es una herramienta diagnóstica crecientemente utilizada para el diagnóstico de TBC. Los predictores de riesgo para TBC pulmonar son variados y varían de acuerdo a la población estudiada. Los objetivos del presente trabajo fueron: evaluar la performance de la detección de M. tuberculosis por la técnica Xpert® MTB/RIF para el diagnóstico de TBC pulmonar y determinar los factores predictores de presencia de esta enfermedad en pacientes asistidos en el Hospital Pasteur de Montevideo. Se realizó un estudio descriptivo, observacional y transversal. Se incluyeron 254 pacientes, 68 con TBC pulmonar. La sensibilidad de la prueba Xpert® MTB/RIF para detectar M. tuberculosis fue 100% (IC 95%: 91,2-100) y la especificidad 95,1% (IC 95%: 83,9-98,7). En el análisis multivariado se evidenció que los predictores independientes para presencia de TBC pulmonar fueron: contacto cercano con otro caso de TBC (p<0,001), consumo de pasta base de cocaína (p=0,006) y presentarse con adelgazamiento (p<0,001). En suma, la prueba Xpert® MTB/RIF se comportó como una excelente herramienta diagnóstica en nuestra población con elevada prevalencia de TBC pulmonar. Los predictores independientes para esta enfermedad indican que en la población analizada las estrategias de control de esta enfermedad requieren un abordaje multidisciplinario.
Summary: According to global estimations, there were approximately 10 million new cases of tuberculosis in 2018. Molecular diagnosis constitutes a rapidly growing diagnostic tool for tuberculosis. Risk predictors for pulmonary tuberculosis are varied and they depend on the population studied. The study aimed to assess the performance of M. tuberculosis detection by use of Xpert® MTB/RIF diagnostic test to diagnose pulmonary tuberculosis and to identify predictive factors for this disease in patients assisted at Pasteur Hospital in Montevideo. A descriptive, observational and transversal study was conducted, which included 254 patients, 68 of which had pulmonary tuberculosis. Sensitivity of the Xpert MTB/RIF assay to detect M. tuberculosis was 100% (CI 95%: 91.2-100) and specificity 95.1% (CI 95%: 83.9-98.7). Multivariate analysis evidenced the following to be the independent predictors that detect pulmonary tuberculosis: close contact with other cases of tuberculosis (p<0.001), coca-paste consumption (p=0.006) and evidence of loss of weight (p<0,001). To sum up, the Xpert® MTB/RIF assay proved to be an excellent diagnostic tool in our population with a high prevalence of pulmonary tuberculosis. Independent predictors for this disease show that, in the population studied, control strategies require a multidisciplinary approach.
Resumo: Globalmente, estima-se que em 2018 ocorreram cerca de 10 milhões de novos casos de tuberculose (TB). A detecção molecular é uma ferramenta diagnóstica cada vez mais usada para seu diagnóstico. Os preditores de risco para TB pulmonar são diversos e variam de acordo com a população estudada. Os objetivos deste estudo foram: avaliar o desempenho da detecção do M. tuberculosis pela técnica Xpert MTB/RIF para o diagnóstico da TB pulmonar e determinar os fatores preditivos da presença desta doença em pacientes atendidos no Hospital Pasteur de Montevidéu. Foi realizado um estudo descritivo, observacional e transversal. 254 pacientes foram incluídos, 68 com TB pulmonar. A sensibilidade do teste Xpert® MTB/RIF para detectar M. tuberculosis foi de 100% (IC 95%: 91,2-100) e a especificidade de 95,1% (IC 95%: 83,9- 98,7). A análise multivariada mostrou que os preditores independentes para a presença de tuberculose pulmonar foram: contato próximo com outro caso de tuberculose (p <0,001), consumo de pasta base de cocaína (p = 0,006) e apresentar perda de peso (p <0,001). Em suma, o teste Xpert® MTB/RIF se comportou como uma excelente ferramenta diagnóstica em nossa população com alta prevalência de TB pulmonar. Os preditores independentes para essa doença indicam que, na população analisada, as estratégias de controle da doença requerem uma abordagem multidisciplinar.
Assuntos
Humanos , Tuberculose Pulmonar/diagnóstico , Técnicas de Diagnóstico Molecular , Mycobacterium tuberculosis , Valor Preditivo dos Testes , Sensibilidade e EspecificidadeRESUMO
OBJECTIVES: The objective of this study was to characterise the mechanisms underlying quinolone and oxyimino-cephalosporin resistance in a Citrobacter freundii clinical isolate obtained from the ICU in a university hospital in Uruguay. METHODS: Citrobacter freundii strain CF638 was isolated from a urine culture. Identification was performed using a VITEK®2 system, and antimicrobial susceptibility was established by MIC determination and disk diffusion assay. Resistance genes and mobile genetic elements were identified by PCR and sequencing. Plasmid transfer was assessed by conjugation and the plasmid size was estimated by S1-PFGE. Plasmid incompatibility (Inc) group and toxin-antitoxin systems were sought by PCR. RESULTS: Strain CF638 showed a multidrug-resistant profile, including resistance to carbapenems and quinolones. Transconjugant TcCF638, harbouring an ca. 200-kb IncA/C plasmid, also showed resistance to all ß-lactams (except aztreonam) and diminished susceptibility to ciprofloxacin. PCR was positive for blaNDM-1 and qnrVC in CF638 and TcCF638. Two different class 1 integrons were detected (In127 and In907). In127 featured the genetic array aadA2-ltr2. Conversely, complex In907 featured two variable regions (VRs); VR-1 consisted of aadB-blaOXA-10-aadA1cc, whereas VR-2 featured a qnrVC6 gene 108bp downstream from ISCR1 and 45bp upstream from qacEΔ1. Expression of qnrVC6 was due to a putative promoter region, detected using the Neural Network Promoter Prediction program. CONCLUSION: To the best of our knowledge, this constitutes the first report of qnrVC within a complex class 1 integron, as well as the first report of the occurrence of such a gene in an NDM-1-producing enterobacterial clinical isolate.