Community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) clones from Paraguayan children.

INTRODUCTION: Staphylococcus aureus is considered one of the most important human pathogens, and its levels of resistance to methicillin have increased even in strains isolated from people without nosocomial risk factors. Molecular analysis is essential for understanding the patterns of dissemination. The objective of this study was to identify community-acquired methicillin-resistant S. aureus (CA-MRSA) clones that infected Paraguayan children patients in two periods of time. METHODOLOGY: An observational, descriptive study was designed to determine the genetic variability of 115 isolates of CA-MRSA recovered from children who attended four reference centers in Paraguay between 2009-2010 and 2012-2013. RESULTS: The combined use of Pulsed Field Gel Electrophoresis (PFGE), Multi-Locus Sequencing Typing, Multi-Locus Variable Analysis (MLVA) and Spa typing techniques allowed the identification of two dominant clones: ST30-IV-t019 (77%) and ST5-IV-t311 (10%), and the establishment of the former as the leading cause of CA-MRSA infections in children during the study period. CONCLUSIONS: This is the first study that provides epidemiological information as well as microbiological and molecular characteristics of CA-MRSA isolates recovered from children from Asunción and the Central Department of Paraguay.

Estandarización del análisis multi-locus de número variable de repeticiones en tándem para el estudio de staphylococcus aureus resistentes a meticilina aislados de la comunidad en Paraguay / Multiple-locus variable-number of tandem repeat analysis standardization for community isolates methicilineresistant staphylococcus aureus study in Paraguay

Staphylococcus aureus es un patógeno capaz de causar infecciones con amplio rango de severidad y adaptarse a diferentes tejidos. Su epidemiología es compleja, por circulación de cientos de clones a nivel mundial, lo que requiere de métodos moleculares reproducibles y de alto poder discriminatorio para la identificación de los mismos. El presente estudio tuvo como objetivo principal la estandarización del análisis multi-locus de número variable de repeticiones en tándem (MLVA) para análisis de variabilidad genética de aislados de S. aureus previamente tipificados por electroforesis en gel de campo pulsado (PFGE), gold standard para tipificación de aislados. La MLVA se realizó por amplificación de 7 locus VNTR (clfA, clfB, sdrC, sdrD, sdrE, sspA y spA) por PCR. Se alcanzó un alto nivel de reproducibilidad. El empleo de cepas previamente tipificadas por análisis de secuencias multi-locus (MLST), PFGE, locus spa y cassette SCCmec, permitió validar de forma comparativa el agrupamiento generado por MLVA. Los aislados que fueron agrupados como idénticos por MLVA presentaron resultados congruentes con la totalidad de las otras técnicas moleculares y esta demostró ser más sensible que PFGE para distinguir entre aislados que presentaron patrones PFGE idénticos. La MLVA cumple todos los criterios de un método de tipificación útil.

Staphylococcus aureus is a pathogen that can produce several infections with a wide range of severity and it has the ability to adapt to different tissues. The epidemiology is complex, due to circulation of many different clones worldwide, so the analysis for its identification requires reproducible and high discriminatory power molecular methods. The aim of this study was to standardize the molecular technique multiple-locus variable number of tandem repeat analysis (MLVA) for the genetic variability analysis of S. aureus isolates, previously characterized by pulsed field gel electrophoresis (PFGE). The MLVA was made by PCR amplification of seven VNTR locus (clfA, clfB, sdrC, sdrD, sdrE, sspA y spA). A high level of reproducibility has been reached in the study. The use of isolates previously typified by multi-locus sequence typing (MLST), PFGE, locus spa and cassette SCCmec, allowed to validate the MLVA clusters comparatively. The isolates that were clustered by MLVA as the same isolate, showed the same results by other molecular techniques, and the MLVA can distinguish isolates with identical PFGE patterns. This technique meets all the criteria of a useful molecular typification technique.

Staphylococcus aureus adquiridos en la comunidad: caracterización clínica, fenotípica y genotípica de aislados en niños paraguayos / Community-acquired Staphylococcus aureus isolated from Paraguayan children: clinical, phenotypic and genotypic characterization

Introduction: The prevalence of Staphylococcus aureus in the community has increased, being the pediatric population the most affected. This fact highlights the need for epidemiological surveillance. Aim: To characterize clinical, phenotypic and genotypic isolates of S. aureus children’s samples with community-acquired infections, collected in hospitals of Asuncion and the Central Department, between November 2009 and December 2010. Materials and Methods: Descriptive and transverse analysis with analytical component. Clinical data collected by medical records, antibiotic susceptibility according to CLSI criteria and detection of mecA (encoding methicillin resistance) and luk-PV genes (encoding Panton Valentine leucocidin) by PCR using specific oligonucleotides. Results: 123 isolates of S. aureus, 76% came from skin and soft tissue infections and 20% from sepsis. 18.7% (n = 23) were resistant to methicillin (MRSA). The presence of the mecA gene, a variant there and the PVL was detected in 12.2 and 48 isolates respectively. 43% of MRSA (n = 10) was carrying luk-PV. The clinical and demographic differences between patients infected with MRSA or MSSA were not statistically significant. Discussion: This study constitutes the first phenotypic and genotypic characterization of S. aureus associated with pediatric patients in Paraguay.

Introducción: La prevalencia de infecciones por Staphylococcus aureus en la comunidad ha aumentado, siendo la población pediátrica la más afectada; poniendo de relieve la necesidad de una vigilancia epidemiológica. Objetivo: Caracterizar clínica, fenotípica y genotípicamente aislados de S. aureus de muestras de niños con infecciones adquiridas en la comunidad, recolectadas en hospitales de Asunción y el Departamento Central, entre noviembre de 2009 y diciembre de 2010. Materiales y Métodos: Estudio descriptivo de corte trasverso. Datos clínicos fueron recabados de fichas, la susceptibilidad a antimicrobianos se hizo según criterio del CLSI y la detección de genes mecA y luk-PV se realizó por RPC empleando oligonucleótidos específicos. Resultados: De 123 aislados de S. aureus, 76% provenían de infecciones de piel y tejidos blandos y 20% de pacientes con bacteriemias. 18,7% (n: 23) fueron resistentes a meticilina (SARM). Se detectó la presencia de genes mecA, una variante del mismo y luk-PV en 9,8%, 1,6 y 39% de los aislados, respectivamente. El 43% de los SARM (n: 10) fue portador de luk-PV. Las diferencias clínicas y demográficas entre pacientes infectados por SARM o SASM no fueron estadísticamente significativas. Discusión: Este estudio constituye la primera caracterización clínica, fenotípica y genotípica de S. aureus asociados a la comunidad en población pediátrica realizada en Paraguay.

[Community-acquired Staphylococcus aureus isolated from Paraguayan children: clinical, phenotypic and genotypic characterization]. / Staphylococcus aureus adquiridos en la comunidad: caracterización clínica, fenotípica y genotípica de aislados en niños paraguayos.

INTRODUCTION: The prevalence of Staphylococcus aureus in the community has increased, being the pediatric population the most affected. This fact highlights the need for epidemiological surveillance. AIM: To characterize clinical, phenotypic and genotypic isolates of S. aureus children's samples with community-acquired infections, collected in hospitals of Asuncion and the Central Department, between November 2009 and December 2010. MATERIALS AND METHODS: Descriptive and transverse analysis with analytical component. Clinical data collected by medical records, antibiotic susceptibility according to CLSI criteria and detection of mecA (encoding methicillin resistance) and luk-PV genes (encoding Panton Valentine leucocidin) by PCR using specific oligonucleotides. RESULTS: 123 isolates of S. aureus, 76% came from skin and soft tissue infections and 20% from sepsis. 18.7% (n = 23) were resistant to methicillin (MRSA). The presence of the mecA gene, a variant there and the PVL was detected in 12.2 and 48 isolates respectively. 43% of MRSA (n = 10) was carrying luk-PV. The clinical and demographic differences between patients infected with MRSA or MSSA were not statistically significant. DISCUSSION: This study constitutes the first phenotypic and genotypic characterization of S. aureus associated with pediatric patients in Paraguay.

Resistencia a antibióticos no betalactámicos de aislamientos invasores de Streptococcus pneumoniae en niños latinoamericanos: SIREVA II, 2000-2005 / Resistance to non-beta-lactam antibiotics in the clinical isolates of Streptococcus pneumoniae of children in Latin America: SIREVA II, 2000-2005

OBJETIVO. Determinar la evolución de la resistencia a la eritromicina, el cloranfenicol, el trimetoprim-sulfametozaxol (SXT) y la vancomicina de aislamientos invasores de Streptococcus pneumoniae obtenidos de niños de 10 países de América Latina y del Caribe en seis años de vigilancia. MÉTODOS. Se analizaron 8 993 aislamientos de S. pneumoniae recuperados entre 2000 y 2005 de niños menores de 6 años con infecciones invasoras, procedentes de Argentina, Brasil, Chile, Colombia, Cuba, México, Paraguay, República Dominicana, Uruguay y Venezuela. La sensibilidad a los antibióticos se determinó mediante los métodos establecidos y estandarizados en el proyecto SIREVA. La resistencia a múltiples antibióticos se definió como la resistencia a tres o más familias de antibióticos, de los no betalactámicos analizados en este estudio o de los betalactámicos evaluados en un estudio previo en el que 37,8% de estos aislamientos presentaron sensibilidad disminuida a la penicilina. RESULTADOS. Se encontró algún grado de resistencia al SXT y la eritromicina (56,4% y 15,4% de los aislamientos estudiados, respectivamente) y 4,6% presentó alta resistencia al cloranfenicol. Todos los aislamientos fueron sensibles a la vancomicina. Se observó la mayor frecuencia de resistencia al SXT en los aislamientos de neumonía y a la eritromicina en los casos de sepsis (61,6% y 25,5%, respectivamente; P < 0,01). La mayor frecuencia de resistencia al SXT se observó en Brasil (71,9%) y a la eritromicina en México (38,2%) y Venezuela (32,9%). Los serotipos 14, 6B, 19F y 23F fueron los que más frecuentemente se asociaron con la resistencia a los antibióticos estudiados. CONCLUSIONES. Se observó una elevada y creciente frecuencia de aislamientos resistentes al SXT y la eritromicina, y una disminución en la proporción de aislamientos resistentes al cloranfenicol. Estas tendencias mostraron diferencias entre los países estudiados.

OBJECTIVE. To examine the development of resistance to erythromycin, chloramphenicol, trimethoprim-sulfamethoxazole (TMP-SMZ), and vancomycin of the invasive isolates of Streptococcus pneumoniae obtained from children in 10 Latin American/Caribbean countries during six years of surveillance. METHODS. Analysis of 8 993 isolates of S. pneumoniae recovered in 2000­2005 from children with invasive infections, who were less than 6 years of age, and from Argentina, Brazil, Chile, Colombia, Cuba, Dominican Republic, Mexico, Paraguay, Uruguay, or Venezuela. Antibiotic susceptibility was determined through the methods established and standardized by the SIREVA project. Multidrug resistance was defined as: resistance to three or more antibiotics of the same class; to the non-beta-lactams analyzed by this study; or, to the beta-lactams evaluated by a previous study, in which 37.8% of these isolates showed decreased susceptibility to penicillin. RESULTS. Some degree of resistance was found to TMP-SMZ and erythromycin (56.4% and 15.4% of the isolates studied, respectively), with 4.6% highly resistant to chloramphenicol. All isolates were susceptible to vancomycin. The highest prevalence of TMP-SMZ resistance was observed in the pneumonia isolates; and that of erythromycin, in cases of sepsis (61.6% and 25.5%, respectively; P < 0.01). The highest prevalence of TMP-SMZ resistance was found in Brazil (71.9%), and that of erythromycin, in Mexico (38.2%) and Venezuela (32.9%). The 14, 6B, 19F, and 23F serotypes were most often associated with resistance to the antibiotics in the study. CONCLUSIONS. High and increasing rates of isolates resistant to TMP-SMZ and erythromycin were observed, as well as a decreasing percentage of isolates resistant to chloramphenicol. These trends highlight differences among the countries studied