B. Alcalá

Development of a Multilocus Sequence Typing Method for Analysis of Listeria monocytogenes Clones

ABSTRACT This study is a first step in the development of multilocus sequence typing (MLST) method for Listeria monocytogenes. Nine housekeeping genes were analyzed in a set of 62 strains isolated from different sources and geographic locations in Spain. These strains were previously characterized by pulsed-field gel electrophoresis (PFGE). Because of low diversity, two loci were discarded from the study. The sequence analysis of the seven remaining genes showed 29 different allelic combinations, with 22 of them represented by only one strain. The results of this sequence analysis were generally consistent with those of PFGE. Because MLST allows the easy comparison and exchange of results obtained in different laboratories, the future application of this new molecular method could be a useful tool for the listeriosis surveillance systems that will allow the identification and distribution of analysis of L. monocytogenes clones in the environment.

Capsule Switching among C:2b:P1.2,5 Meningococcal Epidemic Strains after Mass Immunization Campaign, Spain

A mass immunization campaign for 18-month to 19-year-olds was undertaken in Spain in 1996–1997 because of an epidemic of serogroup C meningococcal disease associated with a C:2b:P1.2,5 strain belonging to the A4 lineage. Surveillance for the “capsule-switching” phenomenon producing B:2b:P1.2,5 isolates was undertaken. Of 2,975 meningococci characterized, B:2b:P1.2,5 and B:2b:P1.2 antigenic combinations were found in 18 isolates; 15 meningococci were defined as serogroup B belonging to the A4 lineage.

Emergence of high level azithromycin-resistant Neisseria gonorrhoeae strain isolated in Argentina.

One Neisseria gonorrhoeae strains highly resistant to azithromycin AzHLR (MIC >2048 mg/L) was isolated in Argentina in 2001 and it has been characterized by N. gonorrhoeae multiantigen sequence typing (NG-MAST) as ST696, suggesting a different event to other isolates in Europe. Neither, mtrR mutations or presence of mef gene were detected.

The epidemic wave of meningococcal disease in Spain in 1996-1997: probably a consequence of strain displacement

During 1996 and 1997 an epidemic wave of meningococcal disease took place in Spain. Initial studies described the antigenic expression of the epidemic strain as C:2b:P1.2,5 and proposed that it was a variant of the previously identified Spanish C:2b:non-subtypable epidemic strain. To clarify this hypothesis, 1036 C:2b:P1.2(5) and 76 C:2b:NST isolates obtained during 1992-1999 were analysed by pulsed-field gel electrophoresis. The majority of the C:2b:P1.2,5 and C:2b:P1.2 isolates showed one of two very closely related profiles. During the epidemic period, 80% of the C:2b:NST strains showed these two pulsotypes. However, before the epidemic wave, most of these C:2b:NST strains (60%) showed a profile that was found infrequently among C:2b:P1.2,5 and C:2b:P1.2 isolates. A similar evolution was observed in C:2b:P1.5 isolates. Thirty-four C:2b:P1.2(5) and 10 C:2b:NST isolates, exhibiting representative pulsotypes, were subjected to multi-locus sequence typing. Isolates belonging to both A4 and ET-37 lineages were identified. These data point to the possibility that the A4 cluster has displaced the ET-37 complex among serogroup C meningococci in Spain.

Sequencing of Neisseria meningitidis penA Gene: the Key to Success in Defining Penicillin G Breakpoints

ABSTRACT Testing of susceptibility to penicillin G by E-test and sequencing of an internal fragment of the penA gene were done for 43 meningococcal strains. Those strains for which the MIC was ≥0.094 μg/ml showed mosaic alleles, so 0.094 μg/ml is suggested as the penicillin G intermediate breakpoint when E-test is used.

Sequencing of the porB gene: a step toward a true characterization of Neisseria meningitidis.

ABSTRACT Variations in class 2/3 (PorB) proteins form the basis for meningococcal serotyping. Antibodies against these proteins are bactericidal, making serotyping results useful not only for epidemiological surveillance of meningococcal disease but also for identifying potential vaccine components. A total of 20 to 60% of meningococcal B and C isolates from any given population are nontypeable (NT) using a panel of monoclonal antibodies. To analyze the mechanisms responsible for the nonserotypeability characteristic in Neisseria meningitidis, we (i) established the nucleotide sequences of porB gene in 146 meningococcal strains (95 not recognized by the serotyping panel), (ii) identified 18 new allelic variants of the porB gene, (iii) correlated allelic variants with serotypes, (iv) suggest the nontypeability characteristic in those 95 NT strains, and (v) reject the possibility of variation in the levels of PorB expression.

First isolate of a Neisseria gonorrhoeae strain associated with an ofloxacin treatment failure in Spain: case report.

FLUOROQUINOLONE REGIMENS, recommended for the treatment of uncomplicated gonorrhea, 1 have been used widely for this purpose. The recent emergence of clinical isolates resistant to fluoroquinolones has become a major problem for the treatment of gonococcal infections in several countries. 2 Genetic characterization of a treatment failure in Spain resulting from a high-level fluoroquinoloneresistant strain of Neisseria gonorrhoeae is reported.

Dynamics of the penA Gene in Serogroup C Meningococcal Strains

The transpeptidase encoding region of the penA gene was sequenced in 44 meningococcal strains (41 serogroup C [23 characterized as serotype 2b and 18 as serotype 2a] and 3 serogroup B [B:2b:P1.2,5]). All strains were characterized by multilocus sequence typing and were determined to be susceptible or intermediate resistant to penicillin (Pen(s) or Pen(i), respectively). A high degree of homology was found among the penA alleles identified in the Pen(s) strains. All the Pen(i) C:2b strains, which belonged to 2 different clonal complexes, showed the same penA gene allele. This fact suggests that 1 of the clonal complexes acquired that allele, spreading it to the other by horizontal transfer. The same allele also was found in the B:2b strains studied, indicating that 1 of the Pen(i) C:2b strains underwent a capsular switching event. A different mosaic penA allele was identified in the Pen(i) C:2a strains, which belonged to the ET37 cluster.

Evolución de la sensibilidad de Neisseria meningitidis a diversos antimicrobianos en el curso de intervenciones con quimioprofilaxis durante un brote epidémico

Introduccion El objetivo del estudio fue evaluar la evolucion de las concentraciones inhibitorias minimas (CIM) poblacionales frente a diversos antimicrobianos en aislamientos de Neisseria meningitidis obtenidos de portadores asintomaticos en el contexto de un brote epidemico de enfermedad meningococica localizado en la provincia de Huelva en el que se realizo una intervencion de quimioprofilaxis en poblacion general. Material y metodos El primer estudio de portadores se realizo en 427 individuos con objeto de conocer la presencia de la cepa causante del brote en poblacion general. Como resultado se procedio a utilizar quimioproflaxis con rifampicina en la poblacion entre 15 y 29 anos (grupo etario con mayor presencia de la cepa epidemica), residentes en la poblacion de Nerva. A los 3 meses se realizo un nuevo estudio de portadores (507 personas) para evaluar el efecto de dicha quimioprofilaxis. Dada la evolucion del brote epidemico fue necesario realizar a los 7 meses una nueva intervencion con ciprofloxacino y transcurridos 2 meses un Nuevo estudio de portadores (399 personas) para evaluar su efecto. Resultados El numero de cepas aisladas en los 3 estudios de portadores realizados fue de 59 (13,8%), 33 (6,5%) y 22 (5,5%), respectivamente. El analisis de la evolucion de la CIM50 y CIM90 de los distintos antibioticos del primer al segundo estudio de portadores (quimioprofilaxis con rifampicina) unicamente detecto cambios con significacion estadistica en las CIM de rifampicina. Asi mismo, cuando se analizo la variacion en la distribucion de las CIM del segundo al tercer estudio (quimioprofilaxis con ciprofloxacino) fueron detectados cambios significativos particularmente en las CIM de cefotaxima. Aunque existieron variaciones de CIM, los porcentajes de sensibilidad al principio y final del estudio no variaron. Conclusiones El empleo de quimioprofilaxis masiva en el grupo de edad con mayor porcentaje de portadores de la cepa de N. meningitidis responsable del brote no modifico sensiblemente la sensibilidad antibiotica de los aislados, si bien puede observarse un aumento de los valores de la CIM50 y CIM90 en el caso de la rifampicina tras la intervencion con este antimicrobiano, y en el caso de la cefotaxima tras las 2 intervenciones realizadas. Las sucesivas intervenciones de quimioprofilaxis con rifampicina y posteriormente con ciprofloxacino tuvieron un claro reflejo en el porcentaje portadores en la poblacion general, con una tendencia claramente decreciente.INTRODUCTION The aim of this study was to assess the evolution of the population MICs for various antimicrobial drugs against Neisseria meningitidis isolates obtained from asymptomatic carriers during a chemoprophylaxis campaign carried out for an epidemic outbreak of meningococcal disease in Nerva, a small town in Huelva province (Spain). MATERIAL AND METHODS A nasopharyngeal carrier study including 427 people was carried out to determine the incidence rate of the epidemic strain among the general population. On the basis of the results, chemoprophylaxis with rifampicin was administered to the population aged 15 to 29 years (age group showing the highest incidence of the epidemic strain among carriers) living in Nerva. Three months later a new carrier study was performed (507 people) to evaluate the effects of chemoprophylaxis. Given the evolution of the outbreak, seven months later a new intervention was required with ciprofloxacin chemoprophylaxis; a second carrier study (399 people) was performed two months later to evaluate its effect. RESULTS The number of strains isolated during the three carrier studies was 59 (13.8%), 33 (6.5%), and 22 (5.5%), respectively. Analysis of the changes in the MIC50 and MIC90 for the various antibiotics from the first to the second carrier study (rifampicin chemoprophylaxis) showed statistical differences only in the distribution of rifampicin MICs. Similarly, when changes from the second to the third study were analyzed (ciprofloxacin chemoprophylaxis), significant variations were detected for the cefotaxime MICs. Nevertheless, although there were changes in the MICs, the percentages of susceptibility from the beginning to the end of the study did not vary. CONCLUSIONS Massive chemoprophylaxis in the age group with the highest incidence of the epidemic strain among carriers did not clearly modify the antibiotic susceptibility of the isolates. However, a slight increase in the MIC50 and MIC90 was observed for rifampicin after the first chemoprophylactic intervention and for cefotaxime at the end of the study. Consecutive chemoprophylactic interventions with rifampicin and ciprofloxacin had an impact on the percentage of meningococcal carriers in the overall population, with a clearly decreasing trend.

Interruption of siaD in a Meningococcal Carrier Isolate Mediated by an Insertion Sequence

Neisseria meningitidis , an important human pathogen causing invasive disease such as meningitis and sepsis, is normally a harmless commensal bacterium that colonizes the human nasopharynx. Dissemination occurs from this ecological niche, which is colonized in up to 10% of the human population ([1][