Antibiotic-resistant Klebsiella pneumoniae and Escherichia coli high-risk clones and an IncFII(k) mosaic plasmid hosting Tn1 (blaTEM-4) in isolates from 1990 to 2004

We describe the genetic background of bla(TEM-4) and the complete sequence of pRYC11::bla(TEM-4), a mosaic plasmid that is highly similar to pKpQIL-like variants, predominant among TEM-4 producers in a Spanish hospital (1990 to 2004), which belong to Klebsiella pneumoniae and Escherichia coli high-risk clones responsible for the current spread of different antibiotic resistance genes. Predominant populations of plasmids and host adapted clonal lineages seem to have greatly contributed to the spread of resistance to extended-spectrum cephalosporins.

Klebsiella pneumoniae sequence type 11 from companion animals bearing ArmA methyltransferase, DHA-1 β-lactamase, and QnrB4

Seven Klebsiella pneumoniae isolates from dogs and cats in Spain were found to be highly resistant to aminoglycosides, and ArmA methyltransferase was responsible for this phenotype. All isolates were typed by multilocus sequence typing (MLST) as ST11, a human epidemic clone reported worldwide and associated with, among others, OXA-48 and NDM carbapenemases. In the seven strains, armA was borne by an IncR plasmid, pB1025, of 50 kb. The isolates were found to coproduce DHA-1 and SHV-11 β-lactamases, as well as the QnrB4 resistance determinant. This first report of the ArmA methyltransferase in pets illustrates their importance as a reservoir for human multidrug-resistant K. pneumoniae.

Polymyxin resistance caused by mgrB inactivation is not associated with significant biological cost in Klebsiella pneumoniae

The inactivation of the mgrB gene, which encodes a negative-feedback regulator of the PhoPQ signaling system, was recently shown to be a common mutational mechanism responsible for acquired polymyxin resistance among carbapenemase-producing Klebsiella pneumoniae strains from clinical sources. In this work, we show that mgrB mutants can easily be selected in vitro from different K. pneumoniae lineages, and mgrB inactivation is not associated with a significant biological cost.

Desarrollo de nuevas formulaciones tópicas de anfotericina B para el tratamiento de micosis

Las infecciones por hongos se han convertido en un tema de gran preocupación en todo el mundo, se estima que más de 40 millones de personas sufren infecciones por hongos, tanto en países desarrollados como en países en vías de desarrollo (Güngör, et al., 2013). Las micosis superficiales se encuentran entre las formas más frecuentes de infecciones en los humanos. Se estima que afectan un 20-25 % de la población mundial y su incidencia está constantemente en aumento (Vena, et al., 2012) (Havlickova, et al., 2008) (Das, et al., 2007). Actualmente, este tipo de infecciones son un frecuente motivo de consulta para el médico de familia (Hernández, et al., 2014) y el dermatólogo. Lo cual nos obliga a permanecer constantemente actualizados La candidiasis es la micosis emergente con mayor efecto en el ser humano debido a su frecuencia y a la gravedad de sus complicaciones (López-Martínez, R., 2010). La candidiasis superficial es una de las formas clínicas más comunes. Es característicamente crónica y recurrente, y, a veces, indica el comienzo de las formas graves de esta micosis (Pappas, et al., 2009). Las levaduras del género Candida son microorganismos pertenecientes a la microbiota normal de individuos sanos, principalmente en la mucosa oral, el tracto gastrointestinal y el tracto genitourinario femenino (Shao, et al., 2007). Sin embargo, estos hongos son responsables de diferentes manifestaciones clínicas, especialmente en pacientes inmunocomprometidos, que van desde infecciones de la piel y mucosas a infecciones sistémicas (Sardi, et al., 2013). Su importancia viene de la alta frecuencia con que colonizan e infectan el huésped humano (De Bernardis, et al., 2004), siendo el cuarto patógeno más común asociado con los casos de infección nosocomial (Wisplinghoff, et al., 2004)...

Evaluación de moxifloxacino como tratamiento alternativo en las infecciones por "Stenotrophomonas maltophilia": optimización de dosis empleando sistemas farmacodinámicos

Stenotrophomonas maltophilia es un patógeno nosocomial, emergente, cuya incidencia en procesos severos esta aumentado de manera análoga al incremento de las poblaciones de pacientes con factores predisponentes. S. maltophilia es un microorganismo ubicuo y adaptado a múltiples ambientes, lo que explica la elevada diversidad genética y el resistoma característico de esta especie que le confiere resistencia a la mayoría de clases de antimicrobianos. Se sugiere que el paciente porta al microorganismo en el momento del ingreso y que la combinación entre la presión antibiótica, especialmente aminoglucósidos y carbapenemas, y el profundo y prolongado estado de inmunosupresión favorecen el desarrollo de la infección. El tratamiento de las infecciones severas, bacteriemias y neumonías, es empírico, y relacionado con una elevada mortalidad, 36-67%, que es atribuida a que la mayoría de pacientes recibe una terapia inapropiada. La carencia de antimicrobianos con suficiente actividad, la rápida aparición de resistencias en los pacientes bajo tratamiento y la ausencia de ensayos clínicos que discriminen tratamientos efectivos, condicionan el complicado manejo de los pacientes infectados por S. maltophilia . En la actualidad solo seis antimicrobianos, cotrimoxazol, ticarcilina-ácido clavulánico, de elección y ceftazidima, levofloxacino, minociclina y cloranfenicol, son considerados tratamientos apropiados para este microorganismo por el CLSI o el EUCAST. La probabilidad de supervivencia aumenta con la rápida administración de agentes sensibles in vitro , pero la mortalidad continúa siendo extraordinariamente alta, 14 y el 45%, lo que demuestra la urgente necesidad de alternativas terapéuticas y de la revaloración de los actuales puntos de corte microbiológicos empleados en el pronóstico de eficacia clínica para esta especie...

Enterococcus faecalis prophage dynamics and contributions to pathogenic traits

Polylysogeny is frequently considered to be the result of an adaptive evolutionary process in which prophages confer fitness and/or virulence factors, thus making them important for evolution of both bacterial populations and infectious diseases. The Enterococcus faecalis V583 isolate belongs to the high-risk clonal complex 2 that is particularly well adapted to the hospital environment. Its genome carries 7 prophage-like elements (V583-pp1 to -pp7), one of which is ubiquitous in the species. In this study, we investigated the activity of the V583 prophages and their contribution to E. faecalis biological traits. We systematically analyzed the ability of each prophage to excise from the bacterial chromosome, to replicate and to package its DNA. We also created a set of E. faecalis isogenic strains that lack from one to all six non-ubiquitous prophages by mimicking natural excision. Our work reveals that prophages of E. faecalis V583 excise from the bacterial chromosome in the presence of a fluoroquinolone, and are able to produce active phage progeny. Intricate interactions between V583 prophages were also unveiled: i) pp7, coined EfCIV583 for E. faecalis chromosomal island of V583, hijacks capsids from helper phage 1, leading to the formation of distinct virions, and ii) pp1, pp3 and pp5 inhibit excision of pp4 and pp6. The hijacking exerted by EfCIV583 on helper phage 1 capsids is the first example of molecular piracy in Gram positive bacteria other than staphylococci. Furthermore, prophages encoding platelet-binding-like proteins were found to be involved in adhesion to human platelets, considered as a first step towards the development of infective endocarditis. Our findings reveal not only a role of E. faecalis V583 prophages in pathogenicity, but also provide an explanation for the correlation between antibiotic usage and E. faecalis success as a nosocomial pathogen, as fluoriquinolone may provoke release of prophages and promote gene dissemination among isolates.

Large-scale screening of a targeted Enterococcus faecalis mutant library identifies envelope fitness factors

Spread of antibiotic resistance among bacteria responsible for nosocomial and community-acquired infections urges for novel therapeutic or prophylactic targets and for innovative pathogen-specific antibacterial compounds. Major challenges are posed by opportunistic pathogens belonging to the low GC% gram-positive bacteria. Among those, Enterococcus faecalis is a leading cause of hospital-acquired infections associated with life-threatening issues and increased hospital costs. To better understand the molecular properties of enterococci that may be required for virulence, and that may explain the emergence of these bacteria in nosocomial infections, we performed the first large-scale functional analysis of E. faecalis V583, the first vancomycin-resistant isolate from a human bloodstream infection. E. faecalis V583 is within the high-risk clonal complex 2 group, which comprises mostly isolates derived from hospital infections worldwide. We conducted broad-range screenings of candidate genes likely involved in host adaptation (e.g., colonization and/or virulence). For this purpose, a library was constructed of targeted insertion mutations in 177 genes encoding putative surface or stress-response factors. Individual mutants were subsequently tested for their i) resistance to oxidative stress, ii) antibiotic resistance, iii) resistance to opsonophagocytosis, iv) adherence to the human colon carcinoma Caco-2 epithelial cells and v) virulence in a surrogate insect model. Our results identified a number of factors that are involved in the interaction between enterococci and their host environments. Their predicted functions highlight the importance of cell envelope glycopolymers in E. faecalis host adaptation. This study provides a valuable genetic database for understanding the steps leading E. faecalis to opportunistic virulence.

Single-nucleotide polymorphism in two representative multidrug-resistant Mycobacterium bovis isolates collected from patients in a Spanish hospital harboring a human infection outbreak

Mycobacterium bovis is the etiological agent of tuberculosis in domestic and wild animals. Its involvement as a human pathogen has been highlighted again with the recent descriptions of transmission through dairy products (18), reactivation or primary infection in human immunodeficiency virus-infected patients (5), and association with meat industry workers, animal keepers, or hunters (3). Strains resistant to antituberculous drugs (M. bovis is naturally resistant to pyrazinamide) pose an additional risk (2). Several studies have demonstrated that mutations in target genes are associated with resistance to antituberculous drugs (4, 7, 10, 11, 16). However, most of them have been developed in Mycobacterium tuberculosis strains and limited data are available regarding M. bovis isolates. The aim of this study was to characterize by sequencing the main genes involved in antibiotic resistance in two multidrug-resistant (MDR) M. bovis isolates in a human outbreak detected in a hospital in Madrid that subsequently spread to several countries (5, 6, 15). The isolates were resistant to 11 drugs, but only their rpoB and katG genes have been analyzed so far (1, 14). We studied the first (93/R1) and last (95/R4) M. bovis isolates of this nosocomial outbreak, characterized by spoligotyping as SB0426 (hexacode 63-5F-5E-7F-FF-60 in the database at www.mbovis.org) (1, 13). Several genes involved in resistance to isoniazid (katG, ahpC, inhA, and the oxyR-ahpC intergenic region), rifampin (rpoB), streptomycin (rrs, rpsL), ethambutol (embB), and quinolones (gyrA) were studied. These genes, or fragments of genes, were amplified and sequenced as previously described (12). The sequence analysis revealed polymorphisms in five (ahpC, rpoB, rpsL, embB, and gyrA) out of nine analyzed genes (Table 1). Nucleotide substitutions in four genes cause a change in the encoded amino acid. Two additional synonymous mutations in ahpC and rpsL differentiated the first and last isolates from the outbreak.

Impacto de sesiones formativas sobre medios de prevención de neumonía asociada a ventilación mecánica en la Unidad de Críticos del Hospital Clínico San Carlos

La neumonía asociada a la ventilación mecánica (NAVM) es la principal infección nosocomial en las Unidades de Cuidados Intensivos. Esta complicación conlleva un aumento de la mortalidad, de los días de estancia y de los costes hospitalarios. El principal mecanismo fisiopatológico es la aspiración repetitiva de secreciones colonizadas de la orofaringe a la vía aérea inferior asociada a procedimientos y productos sanitarios invasivos. Se han estudiado múltiples medidas preventivas publicadas en guías de práctica clínica y revisiones sistemáticas, que utilizadas de manera conjunta, reducen esta infección nosocomial. La correcta aplicación de estas medidas preventivas depende del nivel de conocimiento de los profesionales sanitarios implicados en la asistencia. Las tasas de neumonía en la Unidad Cardiovascular (UCV) del Hospital Clínico San Carlos se mantienen elevadas hasta el año 2009. La formación a los profesionales de enfermería sobre medios de prevención de neumonía asociada al ventilador es una estrategia para mejorar la realización de las intervenciones de estos profesionales y contribuye a la reducción de esta infección nosocomial. El objetivo principal es valorar el impacto de las sesiones formativas a los profesionales de enfermería en el grado de cumplimiento de las intervenciones para prevenir la NAVM (elevación del cabecero, presión del neumotaponamiento, higiene de la orofaringe, colocación y débito de sondaje nasogástrico, tipo de nutrición, humidificación activa, descontaminación digestiva selectiva y aerosolterapia). Los objetivos secundarios son: - Describir el nivel de conocimiento basal de los profesionales de enfermería. - Analizar el impacto de las sesiones formativas en el nivel de conocimiento teórico adquirido. - Determinar la repercusión de las sesiones formativas en la tasa de NAVM, los días de uso de antibiótico, los días de estancia en la UCV y en hospital y la mortalidad...