Antonia Andreu

Molecular characterization of two high-level ceftriaxone-resistant Neisseria gonorrhoeae isolates detected in Catalonia, Spain

OBJECTIVES The aim of this study was to characterize the first two extended-spectrum cephalosporin-resistant and multidrug-resistant (MDR) Neisseria gonorrhoeae isolates collected from two sexually related patients (men who have sex with men) in Spain. METHODS Antimicrobial susceptibility was studied by Etest. Genes involved in quinolone, ceftriaxone and multidrug resistance were amplified by PCR and sequenced in both directions. The isolates were typed by N. gonorrhoeae multi-antigen sequence typing (NG-MAST). RESULTS The two isolates had the same MDR profile, showing resistance to penicillin (MIC 0.094 mg/L; β-lactamase negative), ceftriaxone (MIC 1.5 mg/L), cefixime (MIC 1.5 mg/L), cefotaxime (MIC 1 mg/L), ciprofloxacin (MIC >32 mg/L) and tetracycline (MIC 1.5 mg/L). NG-MAST showed that both isolates belonged to sequence type (ST) 1407 (porB-908 and tbpB-110). Ciprofloxacin resistance was due to amino acid substitutions in GyrA (S91F and D95G) and ParC (S87R). An A deletion in the promoter of the MtrCDE efflux pump (mtrR) was detected. No changes were detected in the pilQ gene. The outer membrane protein PorB showed two substitutions at G120K and A121N. An L421P substitution was observed in the PBP1A (ponA) sequence. The sequence of PBP2 (penA) showed a mosaic structure related to genotype XXXIV with a single additional amino acid substitution (A501P). This genotype was identical to a recently described French isolate (F89). CONCLUSIONS This is the first reported case of high-level extended-spectrum cephalosporin-resistant N. gonorrhoeae transmission. The molecular typing and MDR genotype suggest possible European spread of this strain, highlighting the need for surveillance and the importance of testing the susceptibility of N. gonorrhoeae to extended-spectrum cephalosporins.

Isolation and Characterization of Potentially Pathogenic Antimicrobial-Resistant Escherichia coli Strains from Chicken and Pig Farms in Spain

ABSTRACT To ascertain whether on animal farms there reside extended-spectrum β-lactamase (ESBL) and plasmidic class C β-lactamase-producing Escherichia coli isolates potentially pathogenic for humans, phylogenetic analyses, pulsed-field gel electrophoresis (PFGE) typing, serotyping, and virulence genotyping were performed for 86 isolates from poultry (57 isolates) and pig (29 isolates) farms. E. coli isolates from poultry farms carried genes encoding enzymes of the CTX-M-9 group as well as CMY-2, whereas those from pig farms mainly carried genes encoding CTX-M-1 enzymes. Poultry and pig isolates differed significantly in their phylogenetic group assignments, with phylogroup A predominating in pig isolates and phylogroup D predominating in avian isolates. Among the 86 farm isolates, 23 (26.7%) carried two or more virulence genes typical of extraintestinal pathogenic E. coli (ExPEC). Of these, 20 were isolated from poultry farms and only 3 from pig farms. Ten of the 23 isolates belonged to the classic human ExPEC serotypes O2:H6, O2:HNM, O2:H7, O15:H1, and O25:H4. Despite the high diversity of serotypes and pulsotypes detected among the 86 farm isolates, 13 PFGE clusters were identified. Four of these clusters contained isolates with two or more virulence genes, and two clusters exhibited the classic human ExPEC serotypes O2:HNM (ST10) and O2:H6 (ST115). Although O2:HNM and O2:H6 isolates of human and animal origins differed with respect to their virulence genes and PFGE pulsotypes, the O2:HNM isolates from pigs showed the same sequence type (ST10) as those from humans. The single avian O15:H1 isolate was compared with human clinical isolates of this serotype. Although all were found to belong to phylogroup D and shared the same virulence gene profile, they differed in their sequence types (ST362-avian and ST393-human) and PFGE pulsotypes. Noteworthy was the detection, for the first time, in poultry farms of the clonal groups O25b:H4-ST131-B2, producing CTX-M-9, and O25a-ST648-D, producing CTX-M-32. The virulence genes and PFGE profiles of these two groups were very similar to those of clinical human isolates. While further studies are required to determine the true zoonotic potential of these clonal groups, our results emphasize the zoonotic risk posed especially by poultry farms, but also by pig farms, as reservoirs of ESBL- and CMY-2-encoding E. coli.

Assessment of the Capacity of Lactobacillus to Inhibit the Growth of Uropathogens and Block Their Adhesion to Vaginal Epithelial Cells

To gain insight into the mechanisms by which Lactobacillus blocks the adherence of uropathogens to vaginal epithelial cells and inhibits their growth, 15 Lactobacillus strains and 22 uropathogens were studied. Lactobacilli from hemagglutination group III, identified as Lactobacillus crispatus, showed greater capacity to block uropathogen adherence than those from hemagglutination groups II and I (61.9%, 49.5%, and 52.6% of blockage, respectively). Pseudomonas aeruginosa PA5 and Klebsiella pneumoniae KP7 were the uropathogens most susceptible to blockage, and Staphylococcus aureus SA11 and Proteus mirabilis PM1 were the most resistant. Lactobacillus inhibited uropathogen growth better in liquid assays; the 3 Lactobacillus groups showed similar inhibitory power (72.3%, 71.9%, and 74.2% of light transmittance). P. aeruginosa PA5 was the most inhibited, and Enterococcus species E15 was the least inhibited. There is considerable variation among Lactobacillus strains regarding their adherence to uroepithelium, blockage of uropathogen attachment, and inhibition of uropathogen growth. Although these properties are independent, they may coincide and therefore allow for these strains to balance the vaginal ecosystem and to make them useful as probiotics.

Relationship between Escherichia coli Strains Causing Acute Cystitis in Women and the Fecal E. coli Population of the Host

ABSTRACT Previous epidemiological assessments of the prevalence versus special-pathogenicity hypothesis for urinary tract infection (UTI) pathogenesis in women may have been confounded by underlying host population differences between women with UTI and healthy controls and have not considered the clonal complexity of the fecal Escherichia coli population of the host. In the present study, 42 women with acute uncomplicated cystitis served as their own controls for an analysis of the causative E. coli strain and the concurrent intestinal E. coli population. Clonality among the urine isolate and 30 fecal colonies per subject was assessed by repetitive-element PCR and macrorestriction analysis. Each unique clone underwent PCR-based phylotyping and virulence genotyping. Molecular analysis resolved 109 unique clones (4 urine-only, 38 urine-fecal, and 67 fecal-only clones). Urine clones exhibited a significantly higher prevalence of group B2 than fecal-only clones (69% versus 10%; P < 0.001) and higher aggregate virulence scores (mean, 6.2 versus 2.9; P < 0.001). In multilevel regression models for predicting urine clone status, significant positive predictors included group B2, 10 individual virulence traits, the aggregate virulence score, fecal dominance, relative fecal abundance, and (unique to the present study) a pauciclonal fecal sample. In summary, within the fecal E. coli populations of women with acute cystitis, pauciclonality, clonal dominance, virulence, and group B2 status are closely intertwined. Phylogenetic group B2 status and/or associated virulence factors may promote fecal abundance and pauciclonality, thereby contributing to upstream steps in UTI pathogenesis. This relationship suggests a possible reconciliation of the prevalence and special-pathogenicity hypotheses.

Etiología y sensibilidad a los antimicrobianos de los uropatógenos causantes de la infección urinaria baja adquirida en la comunidad. Estudio nacional multicéntrico

Introduccion El conocimiento de los patrones de sensibilidad a los antimicrobianos es fundamental para orientar el tratamiento empirico y elaborar guias de tratamiento. Se exponen los resultados de un estudio multicentrico que evalua la etiologia y sensibilidad de los principales uropatogenos adquiridos en la comunidad. Metodos Estudio prospectivo realizado entre marzo y julio de 2002, en 15 laboratorios de microbiologia localizados en nueve comunidades autonomas. El urocultivo, las identificaciones bacterianas y las pruebas de sensibilidad se efectuaron en cada laboratorio utilizando la metodologia convencional. Resultados Se obtuvieron 2.724 uropatogenos. El aislado con mayor frecuencia fue Escherichia coli (73%), seguido de Proteus spp. (7,4%), Klebsiella spp. (6,6%) y Enterococcus spp. (4,8%). La sensibilidad de E. coli fue del 97,9% para fosfomicina, del 95,8% para cefixima, del 94,3% para nitrofurantoina, del 90,8% para amoxicilina-acido clavulanico, y del 77,2% para ciprofloxacino. Las resistencias de E. coli a fluoroquinolonas fueron significativamente superiores en varones (28,9% frente a 19% en mujeres; p 32% en Andalucia, Aragon y Castilla y Leon frente a 9,2% en Galicia). Conclusiones E. coli fue el principal agente etiologico. Practicamente todos los aislados de E. coli fueron sensibles a fosfomicina, cefixima y nitrofurantoina. El porcentaje global de resistencia a fluoroquinolonas fue del 23%, aunque vario de manera significativa en funcion de ciertas variables. En Espana, antes de recomendar o instaurar un tratamiento empirico es necesario considerar esta informacion.

Urovirulence determinants in Escherichia coli strains causing prostatitis.

To define the urovirulence properties of Escherichia coli strains producing prostatitis, E. coli strains isolated from men with acute (7 strains) or chronic (23) prostatitis were compared with E. coli isolates from women with pyelonephritis (30), acute cystitis (60), or complicated urinary tract infection (UTI; 30). Strains from prostatitis patients were significantly more likely to express hemolysin than were strains causing complicated UTI (73% vs. 43%; P = .02) and more often demonstrated hybridization with the cytotoxic necrotizing factor-1 (CNF-1) probe (63%) than did strains from women (44%-48%). P fimbrial expression was highest among pyelonephritis (73%) and prostatitis strains (53%) and lowest among E. coli from women with complicated UTI (23%) and cystitis (30%; P < .05, prostatitis strains vs. either of the latter 2 groups). Results suggest that E. coli strains producing prostatitis generally possess urovirulence profiles similar to those of strains from women with acute uncomplicated pyelonephritis and that hemolysin and CNF-1 are especially prevalent in prostatitis strains.

Virulence and antimicrobial resistance profiles among Escherichia coli strains isolated from human and animal wastewater

To gain insight into whether Escherichia coli isolated from humans and resistant to some common antimicrobial agents are derived from animals, 85 E. coli strains were selected by ERIC-PCR from human and animal wastewater samples. Phylogroup, pathogenicity islands (PAIs), resistance to quinolones, fluoroquinolones and presence of extended-spectrum beta-lactamases (ESBLs) were analyzed. Among the total, 55% were resistant to nalidixic acid and 38% to ciprofloxacin; 12% produced ESBLs. Chicken-derived strains were associated with quinolone and fluoroquinolone resistance and presence of ESBLs, while human strains were associated with susceptibility. Group B2 E. coli strains were associated with human origin, susceptibility to fluoroquinolones and presence of PAIs, whereas groups A, B1 and D showed a low virulence profile and a high level of antimicrobial resistance. In both human and animal wastewater, E. coli A, B1 and D were prevalent, and strains from both origins showed a similar virulence profile in each phylogroup. These findings led us to hypothesize that abusive antibiotic use in food animal production may promote the development of resistance among these intestinal E. coli phylogroups, which could later be transmitted to humans through the food supply. The low prevalence of E. coli group B2 in the animal gut may explain, at least in part, the absence of emergence of resistant B2 isolates.

Group B Streptococci Causing Neonatal Infections in Barcelona Are a Stable Clonal Population: 18-Year Surveillance

ABSTRACT We analyzed 212 group B streptococci (GBS) from newborns with invasive infections in the area of Barcelona, Spain, between 1992 and 2009, with the aim of documenting changes in the prevalences of serotypes, antimicrobial resistance, and genetic lineages and evaluating their associations with either early-onset disease (EOD) or late-onset disease (LOD). Serotypes III (n = 118) and Ia (n = 47) together accounted for nearly 78% of the isolates. All isolates carried an alpha or alpha-like protein gene, and specific associations between genes and serotypes, such as serotype Ib and bca, serotype II and bca, serotype III and rib, and serotype V and alp3, reflected the presence of particular genetic lineages. Macrolide resistance (14.2%) was significantly associated with serotype V. Pulsed-field gel electrophoresis (PFGE) clustering was an excellent predictor of serotype and antibiotic resistance. The combination of PFGE and multilocus sequence typing revealed a large number of genetically distinct lineages. Still, specific lineages were dominant in our collection, particularly the serotype III/ST17/rib lineage, which had enhanced potential to cause LOD. Serotype Ia was concentrated in a single PFGE cluster composed of two genetic lineages: ST23/eps and ST24/bca. The ST24/bca sublineage of serotype Ia, which is found infrequently elsewhere, may be emerging as an important cause of neonatal invasive infections in the Mediterranean region. In spite of the introduction of prophylaxis, resulting in a pronounced decline in the frequency of EOD, the study revealed a remarkably stable clonal structure of GBS causing neonatal infections in Barcelona over a period of 18 years.

Papel de Lactobacillus como factor protector de la candidiasis vaginal

Fundamento Esclarecer el papel protector de Lactobacillus frente a la vulvovaginitis candidiasica. Pacientes y metodo Estudio in vitro que determina la capacidad de 15 cepas de Lactobacillus de bloquear la adhesion de dos cepas de Candida albicans y una de C. glabrata a las celulas epiteliales vaginales, asi como la capacidad de inhibir el crecimiento de estas levaduras. Ademas se ha determinado la presencia de Lactobacillus en 115 exudados vaginales de mujeres con vulvovaginitis candidiasica. Resultados Solo 8 de los 15 Lactobacillus bloquearon significativamente la adhesion de C. albicans Y18 a las celulas vaginales. Lactobacillus del grupo hemaglutinante III presentaron la mayor capacidad de bloqueo (62,9%), seguidos de los grupos II (50,6%) y I (26,1%). Ninguno de los 15 Lactobacillus estudiados fue capaz de inhibir en medio solido el crecimiento de las tres levaduras; sin embargo, en medio liquido algunos Lactobacillus evidenciaron cierto poder inhibidor frente a C. albicans Y17 (35,7, 41,7 y 38,1% de transmitancia para los Lactobacillus de los grupos I, II y III, respectivamente). Se detecto la presencia de Lactobacillus en el 89,6% de las mujeres con vulvovaginitis candidiasica, en el 87,8% mediante visualizacion microscopica y en el 72,2% mediante cultivo. Conclusion Los resultados de este estudio sugieren que probablemente Lactobacillus ejerce proteccion contra la vulvovaginitis candidiasica.

Declive de la incidencia de la sepsis perinatal por estreptococo del grupo B (Barcelona 1994-2001). Relación con las políticas profilácticas

Introduccion Analizar la incidencia de la sepsis perinatal por estreptococo del grupo B (EGB) y relacionarla con la aplicacion de las recomendaciones de prevencion consensuadas en 1997 por las sociedades catalanas de Obstetricia, Pediatria y Enfermedades Infecciosas y Microbiologia Clinica. Metodos El estudio se realizo desde 1994 a 2001 y en el participaron 10 hospitales del area de Barcelona, donde se registraron 157.848 nacidos vivos. Resultados Fueron diagnosticados 129 recien nacidos de sepsis perinatal por EGB. La incidencia disminuyo el 86,1%, desde el 1,92/1.000 nacidos vivos en 1994 hasta el 0,26/1.000 en 2001 (p Conclusiones En 8 anos se ha conseguido una importante disminucion de la incidencia de sepsis perinatal por EGB, coincidiendo con la aplicacion de protocolos de prevencion de esta patologia.