Neonatal invasive disease caused by Streptococcus agalactiae in Europe: the DEVANI multi-center study.

PURPOSE: Group B streptococcus (GBS) remains a leading cause of invasive disease, mainly sepsis and meningitis, in infants < 3 months of age and of mortality among neonates. This study, a major component of the European DEVANI project (Design of a Vaccine Against Neonatal Infections) describes clinical and important microbiological characteristics of neonatal GBS diseases. It quantifies the rate of antenatal screening and intrapartum antibiotic prophylaxis among cases and identifies risk factors associated with an adverse outcome. METHODS: Clinical and microbiological data from 153 invasive neonatal cases (82 early-onset [EOD], 71 late-onset disease [LOD] cases) were collected in eight European countries from mid-2008 to end-2010. RESULTS: Respiratory distress was the most frequent clinical sign at onset of EOD, while meningitis is found in > 30% of LOD. The study revealed that 59% of mothers of EOD cases had not received antenatal screening, whilst GBS was detected in 48.5% of screened cases. Meningitis was associated with an adverse outcome in LOD cases, while prematurity and the presence of cardiocirculatory symptoms were associated with an adverse outcome in EOD cases. Capsular-polysaccharide type III was the most frequent in both EOD and LOD cases with regional differences in the clonal complex distribution. CONCLUSIONS: Standardizing recommendations related to neonatal GBS disease and increasing compliance might improve clinical care and the prevention of GBS EOD. But even full adherence to antenatal screening would miss a relevant number of EOD cases, thus, the most promising prophylactic approach against GBS EOD and LOD would be a vaccine for maternal immunization.

Reliable Detection of Group B Streptococcus in the Clinical Laboratory.

Group B streptococcus (GBS) is a leading cause of invasive neonatal infections and a significant pathogen in immunocompromised adults. Screening to detect GBS colonization in pregnant women determines the need for antibiotic prophylaxis in that pregnancy. Efficient determination of the GBS colonization status of pregnant women is crucial. Methods that maximize the probability of GBS recovery are needed. The availability of technologies such as matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS), molecular techniques, and chromogenic culture media, including Granada-type media, have changed the scenario for GBS detection and identification. This review presents and evaluates novel diagnostic tools, as well as classic identification techniques, for GBS species determination.

[Serotypes and antibiotic resistance patterns in beta-hemolytic Streptococcus agalactiae isolates in colonized mothers and newborns with invasive disease]. / Serotipos y patrones de resistencia antibiótica en aislados betahemolíticos de Streptococcus agalactiae de madres colonizadas y recién nacidos con enfermedad invasiva.

INTRODUCTION: Current preventive measures against neonatal disease caused by Streptococcus agalactiae (GBS) are prenatal screening and intrapartum antibiotic prophylaxis with appropriate antimicrobials. An alternative to this strategy would be the administration of a polysaccharide vaccine as the distribution of capsular serotypes of circulating strains needs to be known. METHODS: A study was made of 188 strains from pregnant women carrying GBS and 24 newborns with neonatal disease. Susceptibility testing was performed with penicillin, erythromycin and clindamycin following CLSI standards, and capsular serotype was determined by two methods: latex agglutination and PCR. RESULTS: Of the 188 strains of S.agalactiae from the pregnant women, there was 80.8% agreement in the results between the two techniques. Resistant to erythromycin and clindamycin was found in 16.5% and 10.1%, respectively. For neonatal strains, 95.8% of the results obtained by the two techniques were identical. The rates of resistance to erythromycin and clindamycin were 8.3% and 4.1%, respectively. In both groups, most frequently isolated serotype was iii, and the most related to antimicrobial resistance serotype was v. CONCLUSION: Epidemiological studies are necessary to continue surveillance of serotypes causing invasive disease and its antibiotic sensitivity patterns using sensitive and specific methods.

[Prevention of Neonatal Group B Sreptococcal Infection. Spanish Recommendations. Update 2012. SEIMC/SEGO/SEN/SEQ/SEMFYC Consensus Document]. / Prevención de la infección perinatal por estreptococo del grupo B. Recomendaciones españolas. Actualización 2012. Documento de consenso SEIMC/SEGO/SEN/SEQ/SEMFYC.

Group B streptococci (GBS) remain the most common cause of early onset neonatal sepsis. In 2003 the Spanish Societies of Obstetrics and Gynaecology, Neonatology, Infectious Diseases and Clinical Microbiology, Chemotherapy, and Family and Community Medicine published updated recommendations for the prevention of early onset neonatal GBS infection. It was recommended to study all pregnant women at 35-37 weeks gestation to determine whether they were colonised by GBS, and to administer intrapartum antibiotic prophylaxis (IAP) to all colonised women. There has been a significant reduction in neonatal GBS infection in Spain following the widespread application of IAP. Today most cases of early onset GBS neonatal infection are due to false negative results in detecting GBS, to the lack of communication between laboratories and obstetric units, and to failures in implementing the prevention protocol. In 2010, new recommendations were published by the CDC, and this fact, together with the new knowledge and experience available, has led to the publishing of these new recommendations. The main changes in these revised recommendations include: microbiological methods to identify pregnant GBS carriers and for testing GBS antibiotic sensitivity, and the antibiotics used for IAP are updated; The significance of the presence of GBS in urine, including criteria for the diagnosis of UTI and asymptomatic bacteriuria in pregnancy are clarified; IAP in preterm labour and premature rupture of membranes, and the management of the newborn in relation to GBS carrier status of the mother are also revised. These recommendations are only addressed to the prevention of GBS early neonatal infection, are not effective against late neonatal infection.

Successful restoration of corneal surface integrity with a tissue-engineered allogeneic implant in severe keratitis patients.

OBJECTIVES: Corneal diseases are among the main causes of blindness, with approximately 4.6 and 23 million patients worldwide suffering from bilateral and unilateral corneal blindness, respectively. The standard treatment for severe corneal diseases is corneal transplantation. However, relevant disadvantages, particularly in high-risk conditions, have focused the attention on the search for alternatives. METHODS: We report interim findings of a phase I-II clinical study evaluating the safety and preliminary efficacy of a tissue-engineered corneal substitute composed of a nanostructured fibrin-agarose biocompatible scaffold combined with allogeneic corneal epithelial and stromal cells (NANOULCOR). 5 subjects (5 eyes) suffering from trophic corneal ulcers refractory to conventional treatments, who combined stromal degradation or fibrosis and limbal stem cell deficiency, were included and treated with this allogeneic anterior corneal substitute. RESULTS: The implant completely covered the corneal surface, and ocular surface inflammation decreased following surgery. Only four adverse reactions were registered, and none of them were severe. No detachment, ulcer relapse nor surgical re-interventions were registered after 2 years of follow-up. No signs of graft rejection, local infection or corneal neovascularization were observed either. Efficacy was measured as a significant postoperative improvement in terms of the eye complication grading scales. Anterior segment optical coherence tomography images revealed a more homogeneous and stable ocular surface, with complete scaffold degradation occurring within 3-12 weeks after surgery. CONCLUSIONS: Our findings suggest that the surgical application of this allogeneic anterior human corneal substitute is feasible and safe, showing partial efficacy in the restoration of the corneal surface.

Clarifying the structure of granadaene: total synthesis of related analogue [2]-granadaene and confirmation of its absolute stereochemistry.

Streptococcus agalactiae is an important agent in the infection of neonates in the first world. One of the most extended methods for its identification is based on the detection of a characteristic red pigment in the patient samples, named [12]-granadaene (1). In this article, we present a modular and flexible approach to simple analogues of this ornithine rhamno-polyene 1 and the elucidation of the most important features of its structure: the absolute configuration at C-27, the stereochemistry of the anomeric center and the link of the amino acid ornithine to the rest of the structure.

Molecular characterization and antimicrobial susceptibility of hemolytic Streptococcus agalactiae from post-menopausal women.

PURPOSE: Streptococcus agalactiae (Group B streptococcus, GBS) is increasingly recognized as a pathogen in adult populations, including the elderly. Appropriate treatment involves antibiotics. An alternative to this strategy would be the administration of a polysaccharide vaccine therefore the capsular serotypes and molecular characterization of circulating strains needs to be known. Few studies have been conducted in this population. METHODS: One hundred and seven GBS isolates collected from vagino-rectal swabs from 600 post-menopausal women were analysed for their capsular type, antimicrobial resistance and genetic relatedness (multilocus sequence typing, MLST). RESULTS: The colonization rate was 17.8%. Capsular type III was predominant (34.6%), followed by type V (22.4%). The most frequent sequence type (ST) was 19 (23.3%), followed by 23 (18.7%), 1 (16.8%) and 17 (12.1%). Isolates were assembled into three phylogenetic groups from ST-19, ST-23 and ST-17 founders. All isolates were susceptible to penicillin, whereas resistance to erythromycin and clindamycin was recorded in 23.4% and 20.6% of isolates, respectively. CONCLUSIONS: In our setting, the GBS colonization rate in postmenopausal women is similar to that reported in others populations studied. The population structure of these isolates is highly diverse and contains different STs. These data can contribute to the future development of a polysaccharide vaccine for preventing GBS infection in older adults.

Influenza-like illness criteria were poorly related to laboratory-confirmed influenza in a sentinel surveillance study.

OBJECTIVE: To analyze the most related clinical data for influenza and the utility of influenza-like illness criteria as the clinical threshold for sampling in an influenza sentinel surveillance over a 3-year period. METHODS: Sentinel physicians collected throat specimens and data from outpatients with acute respiratory infection (< or = 72 hours duration). Laboratory-confirmed influenza infection was compared with independent symptoms and the influenza-like illness criteria, as defined by the Classification Committee of the World Organization of Family Doctors. RESULTS: From 1934 patients, 359 (18.56%) yielded positive results for influenza viruses. Only 199 (55.4%) of laboratory-confirmed cases fulfilled clinical criteria of influenza-like illness: positive and negative predictive value (PPV and NPV) of 0.36 and 0.88, respectively. Fever, cough, and rhinorrhea individually correlated with influenza infections (PPV: 0.30, 0.20, and 0.20, respectively; NPV: 0.92, 0.87, and 0.85, respectively). Multivariate analysis demonstrated that the correlation of influenza infection with the presence of fever and cough was similar to the correlation between influenza infection and influenza-like illness criteria (odds ratio 2.24 vs. 2.71, respectively). CONCLUSION: Influenza-like illness criteria are poorly related to laboratory-confirmed influenza. For early detection of influenza viruses in surveillance systems, a less restrictive clinical criterion (specifically, acute respiratory infection) perhaps should be followed.

Non-haemolytic and non-pigmented group b streptococcus, an infrequent cause of early onset neonatal sepsis.

The haemolysin of Group B streptococci (GBS), a leading cause of neonatal infections, is a key virulence factor that has been implicated in the development of invasive infection. The frequency of non-haemolytic (NH) GBS isolates is around 5% among GBS carriers. To determine if similar rates are observed among invasive strains, we evaluated the incidence of NH strains among 199 GBS strains isolated from neonatal blood cultures (first week of life). Overall, we found two (1%) NH strains. This finding suggests that the frequency of NH GBS strains causing early onset invasive neonatal infection is lower than the reported frequency of NH GBS among colonizing strains.

Group B streptococcal haemolysin and pigment, a tale of twins.

Group B streptococcus [(GBS or Streptococcus agalactiae)] is a leading cause of neonatal meningitis and septicaemia. Most clinical isolates express simultaneously a ß-haemolysin/cytolysin and a red polyenic pigment, two phenotypic traits important for GBS identification in medical microbiology. The genetic determinants encoding the GBS haemolysin and pigment have been elucidated and the molecular structure of the pigment has been determined. The cyl operon involved in haemolysin and pigment production is regulated by the major two-component system CovS/R, which coordinates the expression of multiple virulence factors of GBS. Genetic analyses indicated strongly that the haemolysin activity was due to a cytolytic toxin encoded by cylE. However, the biochemical nature of the GBS haemolysin has remained elusive for almost a century because of its instability during purification procedures. Recently, it has been suggested that the haemolytic and cytolytic activity of GBS is due to the ornithine rhamnopolyenic pigment and not to the CylE protein. Here we review and summarize our current knowledge of the genetics, regulation and biochemistry of these twin GBS phenotypic traits, including their functions as GBS virulence factors.

Streptococcus agalactiae: prevención y desarrollo de vacunas / Streptococcus agalactiae: prevention and vaccine development

Streptococcus agalactiae, estreptococo del grupo B (EGB), es la mayor causa de morbi-mortalidad entre los neonatos y un patógeno importante entre los pacientes adultos inmunodeprimidos. A pesar de los avances en la prevención y tratamiento de la infección neonatal, fruto de la implantación de las recomendaciones nacionales e internacionales que en las últimas dos décadas se han desarrollado para ello, aún quedan pendientes mejoras para el control definitivo de la enfermedad. En este sentido, la vacunación frente a EGB podría ser una medida eficaz para la prevención de la infección en aquellos casos donde la profilaxis intraparto no es útil y en pacientes adultos con factores de riesgo de desarrollar infección invasiva por EGB. Esta revisión resume los esfuerzos llevados a cabo para controlar esta infección y aporta información sobre el estado actual de las vacunas frente a EGB empleando diferentes estrategias en su diseño (AU)

Streptococcus agalactiae, group B Streptococcus (SGB), is the most important cause of morbi-mortality among newborn population, and an important pathogen among immunossupressed adult patients. Despite the advances in the treatment and prevention of neonatal infections as a consequence of implementation of national and international recommendations for prevention of infection, there are still some improvements for the final control of the disease. In this sense, the vaccination against SGB could be an effective measure for the prevention of disease in those cases where intrapartum prophylaxis is not useful and in adult patients with risk factors for invasive infection due to SGB. This review summarizes the efforts made until now in order to establish the control of the infection, and brings some information on the current state-of-the art of vaccines against SGB, in which different strategies in their design have been used (AU)

[Prevention of perinatal group B Streptococcal disease. Updated Spanish recommendations 2012]. / Prevención de la infección perinatal por estreptococo del grupo B. Recomendaciones españolas revisadas 2012.

It has been a significant reduction in neonatal group B streptococcus (GBS) infection in Spain following the widespread application of intrapartum antibiotic prophylaxis. In 2010, new recommendations have been published by the CDC and this fact, together with the new knowledge and experience available, has driven to the participating scientific societies publishing these new recommendations. In these recommendations is advised to study all pregnant women at 35-37 gestation weeks` to determine if they are colonized by GBS and to administer intrapartum antibiotic prophylaxis (IAP) to all colonized mothers. Microbiological methods to identify pregnant GBS carriers are updated and intrapartrum antibiotic prophylaxis in preterm labour and premature rupture of membranes and the management of the newborn in relation to GBS carrier status of the mother are also revised.

Serotipos y patrones de resistencia antibiótica en aislados betahemolíticos deStreptococcus agalactiae de madres colonizadas y recién nacidos con enfermedad invasiva / Serotypes and antibiotic resistance patterns in beta-hemolytic Streptococcus agalactiae isolates in colonized mothers and newborns with invasive disease

INTRODUCCIÓN: Las actuales medidas de prevención frente a la enfermedad neonatal causada por Streptococcus agalactiae, estreptococo del grupoB (EGB), son la realización de un cribado prenatal y la administración de profilaxis antibiótica intraparto con antimicrobianos adecuados. Una alternativa a esta estrategia sería la administración de una vacuna polisacarídica, por lo que es necesario conocer la distribución de serotipos capsulares de las cepas circulantes. MÉTODOS: Se estudiaron 188 cepas procedentes de gestantes del área sanitaria norte de Granada portadoras vaginorrectales de EGB y 24 de recién nacidos con enfermedad neonatal enviadas al laboratorio desde distintos hospitales andaluces. Se realizó antibiograma frente a penicilina, eritromicina y clindamicina siguiendo las normas del Clinical and Laboratory Standards Institute (CLSI), y se determinó su serotipo capsular mediante 2 métodos: aglutinación con partículas de látex y métodos moleculares. RESULTADOS: De las 188 cepas de S.agalactiae pertenecientes a mujeres embarazadas, se obtuvo una concordancia en los resultados del 80,8% entre ambas técnicas. Se detectó resistencia a eritromicina y clindamicina en el 16,5 y el 10,1% de cepas, respectivamente. En las cepas neonatales, en el 95,8% de los aislados los resultados obtenidos por ambas técnicas fueron coincidentes. Las tasas de resistencia frente a eritromicina y clindamicina fueron del 8,3 y del 4,1%, respectivamente. En ambos grupos de aislados el serotipo más frecuente fue el III y el más relacionado con resistencia frente a antimicrobianos, el V. CONCLUSIÓN: Se deberían realizar más estudios epidemiológicos que permitan continuar con una vigilancia de los serotipos causantes de enfermedad invasiva así como sus patrones de sensibilidad antibiótica utilizando métodos sensibles y específicos

INTRODUCTION: Current preventive measures against neonatal disease caused by Streptococcus agalactiae (GBS) are prenatal screening and intrapartum antibiotic prophylaxis with appropriate antimicrobials. An alternative to this strategy would be the administration of a polysaccharide vaccine as the distribution of capsular serotypes of circulating strains needs to be known. METHODS: A study was made of 188 strains from pregnant women carrying GBS and 24 newborns with neonatal disease. Susceptibility testing was performed with penicillin, erythromycin and clindamycin following CLSI standards, and capsular serotype was determined by two methods: latex agglutination and PCR. RESULTS: Of the 188 strains of S.agalactiae from the pregnant women, there was 80.8% agreement in the results between the two techniques. Resistant to erythromycin and clindamycin was found in 16.5% and 10.1%, respectively. For neonatal strains, 95.8% of the results obtained by the two techniques were identical. The rates of resistance to erythromycin and clindamycin were 8.3% and 4.1%, respectively. In both groups, most frequently isolated serotype was III, and the most related to antimicrobial resistance serotype was V. CONCLUSIÓN: Epidemiological studies are necessary to continue surveillance of serotypes causing invasive disease and its antibiotic sensitivity patterns using sensitive and specific methods

The putative glycosyltransferase-encoding gene cylJ and the group B Streptococcus (GBS)-specific gene cylK modulate hemolysin production and virulence of GBS.

Group B streptococcus (GBS) expresses a hemolysin/cytolysin that plays an important role in pathogenesis. Using the Himar1 transposon mutagenesis system, a hypohemolytic mutant carrying an interrupted cylJ gene was characterized. cylJ, encoding a putative glycosyltransferase, and cylK, whose product is unknown, are both required for the full hemolytic/cytolytic activity, pigment formation, and virulence of GBS.

Testing of diagnostic methods for detection of influenza virus for optimal performance in the context of an influenza surveillance network.

Influenza surveillance networks must detect early the viruses that will cause the forthcoming annual epidemics and isolate the strains for further characterization. We obtained the highest sensitivity (95.4%) with a diagnostic tool that combined a shell-vial assay and reverse transcription-PCR on cell culture supernatants at 48 h, and indeed, recovered the strain.

[Enterovirus 75, a new pathogenic virus in Granada province (Spain)]. / Enterovirus 75, un nuevo virus patógeno en nuestro medio.

INTRODUCTION: Members of the genus Enterovirus are usually investigated for their etiological role in neurological syndromes. However, they are often associated with other syndromes such as febrile illness, acute respiratory infection and exanthema. In this study, clinical and epidemiological data from five subjects with infection by the recently described enterovirus 75 were analyzed in the province of Granada (Spain). METHODS: Diagnosis at the genus level was carried out by viral culture in MRC-5 and rhabdomyosarcoma cell lines. Isolate serotypes were determined by RT-PCR of a fragment of the VP1 region and subsequent sequencing of the PCR products. RESULTS: Among the five enterovirus 75 isolated, two were detected in children with aseptic meningitis (1 month and 12 years old) and three in subjects with non-neurological syndromes, i.e. acute respiratory infection, febrile illness and gastroenteritis (all were aged less than one year). The five cases were detected between December 2005 and May 2006. All patients recovered without sequelae. CONCLUSION: These data demonstrate that enterovirus 75 circulates in the south of Spain and indicate that this enterovirus serotype may be implicated in less severe non-neurological syndromes, particularly in younger children, and mainly during the cold months of the year.

Granadaene: proposed structure of the group B Streptococcus polyenic pigment.

The goal of this work was to determine the chemical nature of the red pigment produced by Streptococcus agalactiae, which has been thought to be a carotene. We extracted the pigment with 0.1 M KOH and purified it by column chromatography on Sephadex LH. Data from elemental analysis and mass and nuclear magnetic resonance spectra lead us to propose the structure to be that of a new ornithine rhamno-polyene with 12 conjugated double bonds, to which we have assigned the trivial name granadaene.

Specimen storage in transport medium and detection of group B streptococci by culture.

Recovery of group B streptococci (GBS) was assessed in 1,204 vaginorectal swabs stored in Amies transport medium at 4 or 21 degrees C for 1 to 4 days either by direct inoculation onto Granada agar (GA) or by culture in blood agar (BA) and GA after a selective broth enrichment (SBE) step. Following storage at 4 degrees C, GBS detection in GA was not affected after 72 h by either direct inoculation or SBE; however, GBS were not detected after SBE in the BA subculture in some samples after 48 h of storage and in GA after 96 h. After storage at 21 degrees C, loss of GBS-positive results was significant after 48 h by direct inoculation in GA and after 96 h by SBE and BA subculture; some GBS-positive samples were not detected after 24 h of storage followed by SBE and BA subculture or after 48 h of storage followed by SBE and GA subculture. Storage of swabs in transport medium, even at 4 degrees C, produced after 24 h an underestimation of the intensity of GBS colonization in most specimens. These data indicate that viability of GBS is not fully preserved by storage of vaginorectal swabs in Amies transport medium, mainly if they are not stored under refrigeration.