Caracterización molecular de B-Lactamasas de espectro extendido en cepas de escherichia coli causantes de infección urinaria en pacientes inmunocomprometidos / Molecular characterization of extended spectrum B-Lactamases in strains of escherichia coli causing urinary infection in immunocompromised patients

Introducción. La Infección del Tracto Urinario (ITU) es la infección bacteriana que más se diagnostica en el mundo y su agente causal más frecuente es Escherichia coli (E. coli), bacteria que ha adquirido importancia por su capacidad de producir betalactamasa de espectro extendido (BLEE), lo cual dificulta su tratamiento y hace más frecuentes las infecciones recurrentes y recidivantes incluyendo sus complicaciones, sobretodo en pacientes inmunocomprometidos.Objetivo. Identificar pacientes con ITU producidas por E. coli productoras de BLEE y evaluar su espectro antibacteriano y molecular.Material y métodos. Estudio longitudinal y prospectivo realizado con muestras urinarias de 53 pacientes de la Caja Nacional de Salud en La Paz-Bolivia. Se aisló como agente causal a E. coli en el 72% de las muestras, de estas, 35 presentaron fenotipo BLEE sensibles a imipenem, gentamicina y nitrofurantoina (100%) y amikacina (94%). Los ensayos de fenotipificación reportaron predominio del tipo PhP-2 y los filogenéticos (PCR y secuenciación) identificaron predominio de bla CTX-M-15 asociada a bla TEM-1.Conclusión. Los fármacos de primera línea para el tratamiento de la ITU no son adecuados, haciendo a los pacientes más susceptibles a infecciones recurrentes y recidivantes. Se deben identificar precozmente y tratar eficazmente las ITU producidas por cepas de E. coli productoras de BLEE.

Molecular characterization and genetic diversity of ESBL-producing Escherichia coli colonizing the migratory Franklin's gulls (Leucophaeus pipixcan) in Antofagasta, North of Chile.

The role of wild animals, particularly migratory birds, in the dissemination of antibiotic-resistant bacteria between geographically distant ecosystems is usually underestimated. The aim of this work was to characterize the Escherichia coli population from Franklin's gull feces, focusing on the extended-spectrum ß-lactamase (ESBL)-producing strains. In the summer of 2011, 124 fecal swabs from seagulls (1 of each) migrating from the United States and Canada to the coast of Antofagasta, north of Chile, were collected. Samples were seeded on MacConkey agar supplemented with 2 µg/ml of cefotaxime and a single colony from each plate was tested for ESBL production by the double-disk ESBL synergy test. Antibiotic susceptibility was determined by the disk diffusion method and blaESBL genes were amplified and sequenced. The genetic diversity of isolates was explored by pulsed-field gel electrophoresis (PFGE)-XbaI and multilocus sequence typing. A total of 91 E. coli isolates with high rates of antibiotic resistance were identified. Carbapenemase production was not detected, whereas 67 of the 91 (54%) isolates exhibited an ESBL phenotype due to the presence of CTX-M-15 (61.3%), CTX-M-2 (19.3%), CTX-M-22 (16.1%), and CTX-M-3 (1.6%) coding genes. High genetic diversity was observed, with 30 PFGE patterns and 23 sequence types (STs), including ST131 (18%), ST44 (15%), ST617 (9%), and ST10 (9%). Results presented here are complementary to those previously reported by Hernández et al. in the same gull species, but located in the Central Region of Chile. Differences observed between gulls from both areas lead us to hypothesize that gulls from the northern location retain, as gut carriers, those resistant bacteria acquired in the United States and/or Canada.

Molecular characterization and antibiotic resistance of Enterococcus species from gut microbiota of Chilean Altiplano camelids.

BACKGROUND: Enterococcus is one of the major human pathogens able to acquire multiple antibiotic-resistant markers as well as virulence factors which also colonize remote ecosystems, including wild animals. In this work, we characterized the Enterococcus population colonizing the gut of Chilean Altiplano camelids without foreign human contact. MATERIAL AND METHODS: Rectal swabs from 40 llamas and 10 alpacas were seeded in M-Enterococcus agar, and we selected a total of 57 isolates. Species identification was performed by biochemical classical tests, semi-automated WIDER system, mass spectrometry analysis by MALDI-TOF (matrix-assisted laser desorption/ionization with a time-of-flight mass spectrometer), and, finally, nucleotide sequence of internal fragments of the 16S rRNA, rpoB, pheS, and aac(6)-I genes. Genetic diversity was measured by pulsed field gel electrophoresis (PFGE)-SmaI, whereas the antibiotic susceptibility was determined by the WIDER system. Carriage of virulence factors was explored by polymerase chain reaction (PCR). RESULTS: Our results demonstrated that the most prevalent specie was Enterococcus hirae (82%), followed by other non-Enterococcus faecalis and non-Enterococcus faecium species. Some discrepancies were detected among the identification methods used, and the most reliable were the rpoB, pheS, and aac(6)-I nucleotide sequencing. Selected isolates exhibited susceptibility to almost all studied antibiotics, and virulence factors were not detected by PCR. Finally, some predominant clones were characterized by PFGE into a diverse genetic background. CONCLUSION: Enterococcus species from the Chilean camelids' gut microbiota were different from those adapted to humans, and they remained free of antibiotic resistance mechanisms as well as virulence factors.

[Detection of virulence genes in aminoglycoside susceptible and resistant Enterococcus faecalis]. / Detección de genes de virulencia en cepas de Enterococcus faecalis susceptibles y resistentes a aminoglucósidos.

BACKGROUND: Enterococcus spp. is an important cause of nosocomial infections A number of virulence factors that may enhance its ability to colonize have been described. Enterococcus is capable of acquiring resistance genes, including high-level resistance (HLR) to aminoglycoside antibiotics. AIM: to investigate the prevalence of genes encoding virulence factors in aminoglycosides susceptible and resistant E. faecalis. MATERIALS AND METHODS: A total of 80 E. faecalis isolates from clinical (n: 52) and poultry samples (n: 28) were included in this study. Bacterial identification was performed by biochemical tests and phenotypificationwas done using the Phene-PlateTM system. Susceptibility to different antimicrobial agents was determined by the agar dilution method. Virulence genes aceI, agg, gelE and efaA were detected by multiplex PCR. RESULTS: All isolates were susceptible to vancomycin and ampicillin. HLR to gentamicin (13.5%) and streptomycin (9.6%) was detected only in clinical isolates. The phenotyping revealed a great diversity of PhP-types, but only one clone with 7 strains of similar characteristics was found. The efaA gen was detected in 100% of the isolates. aceI gene was present in 94.2% and 75%, agg gene in 73.1%, and 67.9%, and gelE gene in 57.5% and 28.6% of the clinical and chicken isolates, respectively. Only 6 strains with HLR to aminoglycosides, belonging to the same phenotype, had the aceI, agg, gelE and efaA genes. CONCLUSIONS: E. faecalis with virulence genes and HLR to aminoglycosides were isolated from clinical and chicken samples in Antofagasta. More studies will be necessary to establish an association.

Detección de genes de virulencia en cepas de Enterococcus faecalis susceptibles y resistentes a aminoglucósidos / Detection of virulence genes in aminoglycoside susceptible and resistant Enterococcus faecalis

Background: Enterococcus spp. is an important cause of nosocomial infections A number of virulence factors that may enhance its ability to colonize have been described. Enterococcus is capable of acquiring resistance genes, including high-level resistance (HLR) to aminoglycoside antibiotics. Aim: to investigate the prevalence of genes encoding virulence factors in aminoglycosides susceptible and resistant E. faecalis. Materials and Methods: A total of 80 E. faecalis isolates from clinical (n: 52) and poultry samples (n: 28) were included in this study. Bacterial identification was performed by biochemical tests and phenotypificationwas done using the Phene-PlateTM system. Susceptibility to different antimicrobial agents was determined by the agar dilution method. Virulence genes aceI, agg, gelE and efaA were detected by multiplex PCR. Results: All isolates were susceptible to vancomycin and ampicillin. HLR to gentamicin (13.5%) and streptomycin (9.6%) was detected only in clinical isolates. The phenotyping revealed a great diversity of PhP-types, but only one clone with 7 strains of similar characteristics was found. The efaA gen was detected in 100% of the isolates. aceI gene was present in 94.2% and 75%, agg gene in 73.1%, and 67.9%, and gelE gene in 57.5% and 28.6% of the clinical and chicken isolates, respectively. Only 6 strains with HLR to aminoglycosides, belonging to the same phenotype, had the aceI, agg, gelE and efaA genes. Conclusions: E. faecalis with virulence genes and HLR to aminoglycosides were isolated from clinical and chicken samples in Antofagasta. More studies will be necessary to establish an association.

Antecedentes: Enterococcus spp. es una causa importante de infecciones nosocomiales, tanto en Chile como internacional. Se han descrito una serie de factores de virulencia en este microorganismo, que pueden, por ejemplo, aumentar su habilidad para colonizar. Enterococcus tiene capacidad de adquirir genes de resistencia, entre ellos la resistencia de alto nivel (RAN) a los antimicrobianos aminoglucósidos. Objetivo: Investigar la prevalencia de genes de virulencia en cepas de E. faecalis susceptibles y resistentes a aminoglucósidos. Material y Métodos: Un total de 80 cepas de E. faecalis aisladas de muestras clínicas (n: 52) y pollos (n: 28) se incluyeron en este estudio. La identificación se hizo por pruebas bioquímicas y se tipificaron por el sistema Phene-PlateMR. La susceptibilidad a diferentes antimicrobianos fue realizada por test de dilución en agar. Los genes de virulencia aceI, agg, gelE y efaA fueron investigados por RPC múltiple. Resultados: Todas las cepas de E. faecalis fueron susceptibles a vancomicina y ampicilina. Un 13,5% de las cepas clínicas presentaron resistencia de alto nivel a gentamicina y 9,6% a estreptomicina. La tipificación reveló una gran diversidad de fenotipos, pero se encontró un clon con 7 cepas de características similares. El gen efaA estaba presente en 100% de las cepas, gen aceI en 94,2 y 75%, gen agg 73,1 y 67,9% y gen gelE 57,5 y 28,6% de las cepas clínicas y de pollos, respectivamente. Seis cepas con resistencia de alto nivel a aminoglucósidos, que pertenecían a un mismo fenotipo exhibieron los genes efaA, aceI, agg y gelE juntos. Conclusiones: Cepas de E. faecalis que albergan genes de virulencia y con resistencia de alto nivel a aminoglucósidos fueron aisladas de muestras clínicas y de pollos en Antofagasta. Se requieren mayores estudios para establecer una asociación entre estos factores.