Virulence profile of tigecycline-resistant Gram-negative bacilli isolated from river waters using the Caenorhabditis elegans infection model / Perfil de virulência de bacilos Gram-negativos resistentes à tigeciclina isolados das águas dos rios usando o modelo de infecção por Caenorhabditis elegans

Last-resort antibiotics act as ultimate force to overcome multidrug-resistant strains infections. Cases of tigecycline resistance in gram-negative bacilli in clinical settings are reported worldwide, however, there is no data related to tigecycline resistant strains in river water. This study demonstrates seven tigecycline gram-negative bacilli isolated from river water in Rio de Janeiro metropolitan area, their resistance genes, ability of biofilm formation with/without antibiotics and behavior using the nematode Caenohabidits elegans as infection in vivo model. From 24 gram-negative isolated strains, 16 (66.6%) were classified as multidrug-resistant, however, seven (29.1%) presented resistant to all antimicrobial agents tested, including tigecycline and have been identified by MALDI-TOF as A. baumannii, E. aerogenes and P. agglomerans. All tigecycline-resistant strains presented amplification products for ESBL, AME and PMQR and ability of biofilm formation on hydrophilic and hydro­phobic abiotic surfaces with and without antimicrobial agents. The presence of antimicrobials did not inhibit biofilm formation. Tigecycline-resistant strains differed of OP50 control with P<0,0001 indicating its virulence potential, however, none of them were capable to kill all nematodes during 5 days infection. In conclusion, tigecycline-resistant gram-negative strains have important global public health implications due to the therapeutic problems they pose. Further studies and continuous surveillance of tigecycline-resistant strains in both clinical and aquatic environment remains necessary to track and understand the dissemination of tigecycline resistance

Biofilm formation, interaction and survival within A549 pneumocytes of Klebsiella pneumoniae clinical strains: identification of pulsotypes, multidrug-resistance and genes coding for adhesins: Formação de biofilme, interação e sobrevivência dentro dos pneumócitos A549 de cepas clínicas de Klebsiella pneumoniae: identificação de pulsótipos, multirresistência a drogas e codificação de genes para adesinas

Klebsiella pneumoniaehas become one of the major causes of hospital-acquired infections over decades due to the spread of virulent clones harboring resistant genes to multiple antimicrobial agents. The aim of this study was to investigate phenotypic and genotypic features of virulence mechanism expressed by K. pneumoniae clinical isolates of different PFGE types, including biofilm formation, interaction with pneumocytes A549 lineage and experimental infection by using C. elegans nematodes.  A total of 17 K. pneumoniae strains were isolated from different clinical specimens including blood, urine and respiratory infections. In this present study, 11 strains presented a varied multidrug-resistance profile harboring resistance genes coding for betalactams, aminoglicosydes, fluorquinolones and carbapenemases. PFGE analysis demonstrated the presence of four distinct pulsotypes among K. pneumoniae strains harboring virulence genes for siderophores and fimbiae type 1 and type 3. High adherence and biofilm formation were positively correlated for both polystyrene and glass surfaces in all K. pneumoniae strains analyzed. K. pneumoniae clinical strains showed the ability of adherence, internalization and persistence within human pulmonary epithelial A549 cell line, at different levels. Respiratory infections demonstrated a higher heterogeneity of PFGE types and levels of adherence, intracellular survival and persistence.K. pneumoniae strains were also submitted to Carnohabidits elegans in vivo infection model and data showed that after 24 hr almost 10% of urine-culture isolates worms were dead evidencing virulence profile. Notably, K. pneumoniae strains, presenting virulence genes, was significantly more virulent than those who did not presented any virulence gene after 5 days (survival >60% and >40%)

Detection of bacterial samples on the aquatic ecosystems adjacent to Saquarema Lagoon – Rio de Janeiro

Introduction: Saquarema Lagoon (RJ) has a high ecological and economic value owing to its multiple uses. The population’s constant growth increases the amount of sewage containing bacteria and antimicrobial drugs that are discharged to the environment. Objectives: to detect Gram negative bacilli able to colonize or infect humans and animals and determine their antimicrobial resistance profiles. Methodology:samples were collected in the city centre in April 2010 and at Jaconé (Lagoon’s most preserved site) in February 2011. The total and thermo tolerant coliforms were determined and the isolation of samples was made using agar media containing  32cg/ mL of cephalotin. All samples were tested for antimicrobial susceptibility (AST) and on 16 samples, plasmid DNA was extracted. Results: different Gram negative bacteria were detected, such as: Enterobacter spp, Citrobacter freundii, Klebsiella pneumoniae and Pseudomonas aeruginosa. The coliform results showed that the water quality is proper for Human recreation. AST results demonstrated the existence of bacteria resistant to antimicrobial drugs frequently used in the community. It was possible to detected high molecular weight plasmids and nine samples (56,25%) showed at least one plasmid DNA electrophoresis band. Conclusions: there were not detected resistant samples to antimicrobial drugs normally used in hospital settings, which may possibly refute the idea of a contamination by nosocomial and/or veterinary sewage discharge

Detección de enterobacterias multirresistentes aisladas en aguas de los ríos que desembocan en la bahía de Guanabara y en muestras de hospitales de Río de Janeiro, Brasil

Introduction: The use of antibiotics in humans, animal husbandry and veterinary activities induces selective pressure leading to the colonization and infection by resistant strains.Objective: We evaluated water samples collected from rivers of the Guanabara Bay, which have suffered minor and major environmental degradation, and clinical samples of hospital origin to detect evidence of the presence of resistance genes to aminoglycosides, beta-lactam antibiotics and fluoroquinolones in strains of Klebsiella pneumoniae subsp. pneumoniae, K. pneumoniae subsp. ozaenae and Escherichia coli.Materials and methods: For isolation of the water strains we employed culture media containing 32 μg/ml cephalotin and 8 μg/ml gentamicin. The strains from clinical materials were selected using culture media containing 8 μg/ml gentamicin. The strains were identified and subjected to antimicrobial susceptibility testing (AST), plasmid DNA extraction and polymerase chain reaction (PCR) to detect genes encoding enzymes modifying aminoglycosides (EMA), extended-spectrum beta-lactamases (ESBL) and plasmid mechanisms of quinolone resistance (PMQR).Results: The AST of the isolates recovered from water samples showed multidrugresistance profiles similar to those found in isolates recovered from clinical materials. All isolates from water samples and 90% of the isolates from clinical samples showed at least one plasmid band. In the PCR assays, 7.4% of the isolates recovered from water samples and 20% of those from clinical materials showed amplification products for the three antimicrobial classes.Conclusion: We believe that the detection of microorganisms presenting genetic elements in environments such as water is necessary for the prevention and control of their dissemination with potential to infect humans and other animals in eventual contact with these environments.Introducción. El uso de antibióticos en seres humanos, en la industria pecuaria y en las actividades veterinarias induce una presión selectiva que resulta en la colonización e infección con cepas resistentes.Objetivo. Determinar la presencia de genes de resistencia a aminoglucósidos, betalactámicos y fluoroquinolonas en cepas de Klebsiella pneumoniae subsp. pneumoniae, K. pneumoniae subsp. ozaenae y Escherichia coli, obtenidas de muestras de agua de los ríos que desembocan en la bahía de Guanabara y de muestras clínicas de hospitales de Río de Janeiro. Materiales y métodos. En la selección de las cepas resistentes obtenidas de las muestras de agua de los ríos, se emplearon medios de cultivo que contenían 32 μg/ml de cefalotina y 8 μg/ml de gentamicina. En el caso de las muestras de especímenes clínicos, se usaron medios de cultivo que contenían 8 μg/ml de gentamicina. Las cepas se identificaron y se sometieron a pruebas de sensibilidad antimicrobiana, extracción de ADN plasmídico y pruebas de reacción en cadena de la polimerasa (PCR) para detectar los genes que codifican aquellas enzimas que modifican los aminoglucósidos, las betalactamasas de espectro extendido (BLEE) y los mecanismos de resistencia a las quinolonas mediados por plásmidos.Resultados. Se encontraron perfiles de resistencia a los antimicrobianos similares en los dos grupos. En todas las bacterias obtenidas de las muestras de agua y en 90 % de las muestras clínicas, se evidenciaron bandas de plásmidos asociados con la transferencia de genes de resistencia. En las pruebas de PCR, se obtuvieron productos de amplificación de los genes de resistencia para las tres clases de antimicrobianos analizados, en el 7,4 % de las bacterias recuperadas de las muestras de agua y en el 20 % de aquellas recuperadas de las muestras clínicas.Conclusión. La detección de microorganismos con elementos genéticos que confieren resistencia a los antibióticos en ambientes como el agua, es una estrategia necesaria para prevenir y controlar la diseminación de estos agentes patógenos con potencial para infectar a humanos y a otros animales en dichos ambientes

RESISTÊNCIA ANTIMICROBIANA EM STAPHYLOCOCCUS CAPITIS ISOLADO DE HEMOCULTURA: CARACTERIZAÇÃO FENOTÍPICA E ANÁLISES GENÔMICAS

Introdução/Objetivo: Os estafilococos coagulase-negativa (SCoN), membros da microbiota residente da pele humana, apresentam potencial de causar infecções oportunistas, sobretudo quando ocorre o rompimento da barreira cutânea, seja por trauma ou pela introdução de dispositivos médicos. Dentre os SCoN, Staphylococcus capitis destaca-se como uma das espécies mais frequentes em infecções relacionadas à assistência à saúde (IRAS), principalmente no ambiente hospitalar. A letalidade das infecções está diretamente associada à expressão de fatores de virulência e resistência aos agentes antimicrobianos pelo microrganismo. Este trabalho teve por objetivo investigar a resistência antimicrobiana em uma cepa de Staphylococcus capitis subsp. urealyticus isolada de hemocultura através da determinação do perfil de susceptibilidade aos antimicrobianos e da busca por genes de resistência no genoma completamente sequenciado. Métodos: Foi utilizada uma cepa de Staphylococcus capitis oriunda de hemocultura de um indivíduo adulto e previamente identificada por espectrometria de massas MALDI-TOF. O perfil de susceptibilidade aos antimicrobianos foi determinado de acordo com o BrCAST (2023). O genoma da cepa foi extraído, purificado e completamente sequenciado na plataforma NextSeq 550 (Illumina®). A confirmação da espécie foi realizada pela análise de sequência multilocus (MLSA) e a busca por genes de resistência antimicrobiana foi realizada com a ferramenta de bioinformática ResFinder 4.1. Resultados: A cepa de S. capitis subsp. urealyticus exibiu um fenótipo multidroga-resistente (MDR), apresentando resistência à oxacilina, cefoxitina, norfloxacino, entre outros. A cepa foi identificada como S. capitis subsp. urealyticus pela MLSA. Genes que conferem resistência a diversas classes de antimicrobianos, dentre os quais macrolídeos, aminoglicosídeos, β-lactâmicos e lincosamidas, foram encontrados no genoma sequenciado. Conclusão: As análises revelaram a presença de diversos genes de resistência antimicrobiana no genoma da cepa de S. capitis subsp. urealyticus isolada de hemocultura, corroborando o fenótipo MDR exibido. Estes resultados enfatizam a importância de investigar o perfil de susceptibilidade aos antimicrobianos de isolados clínicos de SCoN, bem como alertam para a necessidade de alternativas para o tratamento das IRAS por microrganismos destas espécies expressando perfis de MDR