Streptococcus pneumoniae serotype 19A in hospitalized children with invasive pneumococcal disease after the introduction of conjugated vaccines in Lima, Peru.

BACKGROUND: The Pneumococcal conjugate vaccine (PCV) has decreased cases of invasive pneumococcal disease (IPD) worldwide. However, the impact of PCVs introduction may be affected by the serotype distribution in a specific context. METHODS: Cross-sectional multicenter passive surveillance study of IPD cases in pediatric patients hospitalized in Lima, Peru between 2016 and 2019 (after PCV13 introduction) to determine the serotype distribution and antimicrobial resistance of Streptococcus pneumoniae. Serotyping was performed by a sequential multiplex PCR and confirmed by whole genome sequencing. RESULTS: Eighty-five S. pneumoniae isolates were recovered (4.07/100,000 among children <60 months of age). Serotype 19A was the most common (49.4%). Children infected with serotype 19A in comparison with children infected with other serotypes were younger, had a lower rate of meningitis and higher rates of pneumonia, complicated pneumonia and antimicrobial resistance; 28.6% of patients with serotype 19A have received at least one dose of PCV13 vs. 62.8% of patients with other serotypes. Using MIC-breakpoints, 81.2% (56/69) of non-meningitis strains and 31.2% (5/16) of meningitis strains were susceptible to penicillin; 18.8% (3/16) of meningitis strains had intermediate resistance to ceftriaxone. Resistance to azithromycin was 78.8% (67/85). Serotype 19A frequency increased over time in the same study population, from 4.2% (4/96) in 2006-2008, to 8.6% (5/58) in 2009-2011, to 49.4% (42/85) in the current study (2016-2019) (p < 0.001). CONCLUSIONS: After PCV13 introduction in Peru, serotype 19A remains the most prevalent; however, the vaccination coverage is still not optimal. Therefore, additonal surveillance studies are needed to determine the remaining IPD burden.

Pneumococcal serotypes and antibiotic resistance in healthy carriage children after introduction of PCV13 in Lima, Peru.

OBJECTIVE: To determinate the frequency of Streptococcus pneumoniae nasopharyngeal carriers, serotypes and antimicrobial resistance in healthy children in Lima, Peru, post-PCV13 introduction and to compare the results with a similar study conducted between 2006 and 2008 before PCV7 introduction (pre-PCV7). METHODS: A cross-sectional multicenter study was conducted between January 2018 and August 2019 in 1000 healthy children under two years of age. We use standard microbiological methods to determinate S. pneumoniae from nasopharyngeal swab, Kirby Bauer and minimum inhibitory concentration methods to determinate antimicrobial susceptibility and whole genomic sequencing to determinate pneumococcal serotypes. RESULTS: The pneumococcal carriage rate was 20.8 % vs. 31.1 % in pre-PCV7 (p < 0.001). The most frequent serotypes were 15C, 19A and 6C (12.4 %, 10.9 % and 10.9 % respectively). The carriage of PCV13 serotypes after PCV13 introduction decreased from 59.1 % (before PCV7 introduction) to 18.7 % (p < 0.001). Penicillin resistance was 75.5 %, TMP/SMX 75.5 % and azithromycin 50.0 %, using disk diffusion. Penicillin resistance rates using MIC breakpoint for meningitis (MIC ≥ 0.12) increased from 60.4 % to 74.5 % (p = 0.001). CONCLUSION: The introduction of PCV13 in the immunization program in Peru has decreased the pneumococcal nasopharyngeal carriage and the frequency of PCV13 serotypes; however, there has been an increase in non-PCV13 serotypes and antimicrobial resistance.

Increase of Macrolide-Resistance in Streptococcus pneumoniae Strains After the Introduction of the 13-Valent Pneumococcal Conjugate Vaccine in Lima, Peru.

Streptococcus pneumoniae upper respiratory infections and pneumonia are often treated with macrolides, but recently macrolide resistance is becoming an increasingly important problem. The 13-valent pneumococcal conjugate vaccine (PCV13) was introduced in the National Immunization Program of Peru in 2015. This study aimed to evaluate the temporal evolution of macrolide resistance in S. pneumoniae isolates collected in five cross-sectional studies conducted before and after this vaccine introduction, from 2006 to 2019 in Lima, Peru. A total of 521 and 242 S. pneumoniae isolates recovered from nasopharyngeal swabs from healthy carrier children < 2 years old (2 carriage studies) and samples from normally sterile body areas from pediatric patients with invasive pneumococcal disease (IPD) (3 IPD studies), respectively, were included in this study. Phenotypic macrolide resistance was detected using the Kirby-Bauer method and/or MIC test. We found a significant increase in macrolide resistance over time, from 33.5% to 50.0% in carriage studies, and from 24.8% to 37.5% and 70.8% in IPD studies. Macrolide resistance genes [erm(B) and mef(A/E)] were screened using PCR. In carriage studies, we detected a significant decrease in the frequency of mef(A/E) genes among macrolide-resistant S. pneumoniae strains (from 66.7% to 50.0%) after introduction of PCV13. The most common mechanism of macrolide-resistant among IPD strains was the presence of erm(B) (96.0%, 95.2% and 85.1% in the 3 IPD studies respectively). Macrolide resistance was more common in serotype 19A strains (80% and 90% among carriage and IPD strains, respectively) vs. non-serotype 19A (35.5% and 34.4% among carriage and IPD strains, respectively). In conclusion, S. pneumoniae macrolide resistance rates are very high among Peruvian children. Future studies are needed in order to evaluate macrolide resistance trends among pneumococcal strains, especially now after the COVID-19 pandemic, since azithromycin was vastly used as empiric treatment of COVID-19 in Peru.

Pathogenicity Island O-122 in enteropathogenic Escherichia coli strains is associated with diarrhea severity in children from Lima Peru.

EPEC is an attaching and effacing diarrheal pathogen that carries a large pathogenicity island, locus for enterocyte effacement (LEE). Recently, the pathogenicity island PAI O-122 was described among non-LEE effectors and found to be associated with diarrhea among atypical EPEC strains. It is unknown if incomplete PAI O-122 could be associated with diarrhea duration and severity. To identify these virulence determinants we analyzed 379 EPEC strains isolated from Peruvian children. EPEC was diagnosed by PCR(eae+, stx-) and classified as typical(t-EPEC) or atypical(a-EPEC). To characterize PAI O-122 we amplified three modules by PCR: Module 1(pagC), Module 2(senA, nleB and nleE) and Module 3(lifA/efa-1). To characterize the large ORF lifA/efa-1 we amplified the regions known as efa-N, efa-M and efa-C. Clinical information was obtained from the cohort study. A total of 379 EPEC strains were able to analyze PAI O-122 genes, 128 (10.4%) EPEC strains were isolated from 1235 diarrhea episodes and 251(9.2%) from 2734 healthy controls. t-EPEC strains were isolated from 14.8% (19/128) of children with diarrhea and 25/251(10.0%) from healthy controls. The most frequent PAI O-122 genes were nleE(37.7%), senA(34.6%) and nleB(37.5%), with similar prevalence among diarrhea and control samples. However, lifA/efa-1 was more common among diarrhea cases than healthy control cases (30.5% vs. 21.1%, p<0.05). The presence of complete PAI O-122 was associated with diarrhea episodes of higher severity among single pathogen infection (33.3% vs. 1.8%, p<0.05) mainly due to the presence of a complete lifA/efa-1 gene. In summary, the gene lifA/efa-1 is significantly associated with diarrheal episodes of higher severity, suggesting to be an important virulent factor.

Resistencia antibiótica de streptococcus pneumoniae en portadores nasofaríngeos sanos de siete regiones del Perú / Antibiotic resistance of streptococcus pneumoniae among healthy nasopharyngeal carriers in seven regions of Peru

Objetivos. Determinar el patrón de susceptibilidad antibiótica de cepas de Streptococcus pneumoniae aisladas de portadores nasofaríngeos sanos menores de 2 años de siete regiones del Perú. Materiales y métodos. Entre el 2007 y 2009 se tomaron muestras de hisopado nasofaríngeo a 2123 niños sanos entre 2 y 24 meses de edad en los consultorios de crecimiento y desarrollo (CRED) y vacunación de hospitales y centros de salud de Lima, Piura, Cusco, Abancay, Arequipa, Huancayo, e Iquitos. Se determinó la resistencia a diez antibióticos mediante la prueba de disco-difusión de las cepas de neumococo aisladas. Resultados. Se aislaron 572 cepas. Se encontró altas tasas de resistencia a cotrimoxazol (58%); penicilina (52,2% no-sensibles); tetraciclina (29,1%); azitromicina (28,9%), y eritromicina (26,3%). La resistencia a cloranfenicol fue baja (8,8%). Se encontró 29,5% de multirresistencia. La resistencia a la azitromicina y a la penicilina fue diferente en las siete regiones (p<0,05), hallándose el mayor porcentaje de cepas no-sensibles a penicilina en Arequipa (63,6%), mientras que el menor fue en Cusco (23,4%). Conclusiones. Los elevados niveles de resistencia encontrados para penicilina, cotrimoxazol y macrólidos en cepas de neumococo aisladas de portadores sanos en todas las regiones estudiadas, y su asociación con uso previo de antibióticos, representan un importante problema de salud pública en nuestro país. Esto resalta la necesidad de implementar, a nivel nacional, estrategias para disminuir el uso irracional de antibióticos, sobre todo en la población pediátrica. Es necesario complementar los datos de resistencia a penicilina con la determinación de la concentración mínima inhibitoria para hacer las recomendaciones terapéuticas respectivas.

Objectives. To determine the pattern of antibiotic susceptibility of isolated Streptococcus pneumoniae strains of healthy nasopharyngeal carriers younger than 2 years in seven regions of Peru. Materials and methods. Between 2007 and 2009, nasopharyngeal swab samples were collected among 2123 healthy children aged 2-24 months in growth and development medical practices (CRED) and vaccination offices of hospitals and health centers in Lima, Piura, Cusco, Abancay, Arequipa, Huancayo, and Iquitos. The resistance to ten antibiotics through disk diffusion sensitivity testing of isolated pneumococcus strains was determined. Results. 572 strains were isolated. High rates of resistance to co-trimoxazole (58%), penicillin (52.2% non-sensitive); tetracycline (29,1%); azithromycin (28,9%), and erythromycin (26,3%). Resistance to chloramphenicol was low (8.8%). Multiresistance was found at 29.5%. Resistance to azithromycin and penicillin was different in all seven regions (p<0,05), the highest percentage of non-sensitive strains being found in Arequipa (63,6%), whereas the lowest percentage was found in Cusco (23.4%). Conclusions. High levels of resistance found to penicillin, co-trimoxasole and macrolides in isolated pneumococcus strains of healthy carriers in all studied regions, and their association to a previous use of antibiotics, represent a significant public health problem in our country. This emphasizes the need to implement nationwide strategies to reduce the irrational use of antibiotics, especially among children. It is necessary to complement data of resistance to penicillin with the determination of minimal inhibitory concentration to make proper therapeutic recommendations.

[Antibiotic resistance of streptococcus pneumoniae among healthy nasopharyngeal carriers in seven regions of Peru]. / Resistencia antibiótica de streptococcus pneumoniae en portadores nasofaríngeos sanos de siete regiones del Perú

OBJECTIVES: To determine the pattern of antibiotic susceptibility of isolated Streptococcus pneumoniae strains of healthy nasopharyngeal carriers younger than 2 years in seven regions of Peru. MATERIALS AND METHODS: Between 2007 and 2009, nasopharyngeal swab samples were collected among 2123 healthy children aged 2-24 months in growth and development medical practices (CRED) and vaccination offices of hospitals and health centers in Lima, Piura, Cusco, Abancay, Arequipa, Huancayo, and Iquitos. The resistance to ten antibiotics through disk diffusion sensitivity testing of isolated pneumococcus strains was determined. RESULTS: 572 strains were isolated. High rates of resistance to co-trimoxazole (58%), penicillin (52.2% non-sensitive); tetracycline (29,1%); azithromycin (28,9%), and erythromycin (26,3%). Resistance to chloramphenicol was low (8.8%). Multiresistance was found at 29.5%. Resistance to azithromycin and penicillin was different in all seven regions (p<0,05), the highest percentage of non-sensitive strains being found in Arequipa (63,6%), whereas the lowest percentage was found in Cusco (23.4%). CONCLUSIONS: High levels of resistance found to penicillin, co-trimoxasole and macrolides in isolated pneumococcus strains of healthy carriers in all studied regions, and their association to a previous use of antibiotics, represent a significant public health problem in our country. This emphasizes the need to implement nationwide strategies to reduce the irrational use of antibiotics, especially among children. It is necessary to complement data of resistance to penicillin with the determination of minimal inhibitory concentration to make proper therapeutic recommendations.

[Pneumococcal serotypes in carrier children prior to the introduction of anti-pneumococcal vaccines in Peru]. / Serotipos de neumococo en niños portadores antes de la vacunación antineumocócica en el Perú.

OBJECTIVES: To determine the carriage rate and serotype distribution of Streptococcus pneumoniae in the nasopharynx of healthy children younger than 2 years prior to the universal use of the pneumococcal conjugate vaccines in Peru. MATERIALS AND METHODS: Between 2007 and 2009 we collected nasopharyngeal swab samples from 2,123 healthy children aged 2 to 24 months in the vaccination and healthy children consultation offices of pediatric hospitals and health centers in 7 cities in Peru: on the coast (Lima, Piura), highlands (Cusco, Abancay, Arequipa and Huancayo) and amazon basin (Iquitos). The pneumococcal strains were isolated and identified at the central laboratory of the project in Lima, and serotyped by Quellung reaction in the pneumococcal reference laboratory at the Center for Diseases Control and Prevention (CDC). RESULTS: We found 27% (573/2123) of pneumococcal nasopharyngeal healthy carrier children. Among the 526 analyzed strains, we found 42 serotypes; the most common were: 19F (18.1%), 6B (14.3%); 23F (8.9%) and 14 (6.5%). CONCLUSIONS: The distribution of vaccine serotypes in the analyzed strains was of 50% for the serotypes present in the seven-valent vaccine, 50.2% for the serotypes present in the ten-valent vaccine and 57.2% for those present in the thirteen-valent vaccine.

Serotipos de neumococo en niños portadores antes de la vacunación antineumocócica en el Perú / Pneumococcal serotypes in carrier children prior to the introduction of anti-pneumococcal vaccines in Peru

Objetivos. Determinar la frecuencia y distribución de serotipos de S. pneumoniae en portadores nasofaríngeos sanos menores de dos años previa al uso universal de la vacuna conjugada antineumocócica en el Perú. Materiales y métodos. Entre los años 2007 y 2009 se tomaron muestras de hisopado nasofaríngeo a 2123 niños sanos entre 2 y 24 meses de edad en los consultorios de crecimiento y desarrollo o vacunación de hospitales y centros de salud de siete ciudades del Perú: costa (Lima, Piura); sierra (Cusco, Abancay, Arequipa y Huancayo) y selva (Iquitos). Las cepas de neumococo fueron aisladas e identificadas en el laboratorio central del proyecto en Lima y serotipificadas por reacción de Quellung en el Laboratorio de Referencia de Neumococo del Centro de Control y Prevención de Enfermedades. Resultados. Se encontró 27,0% (573/2123) de portadores nasofaríngeos sanos de neumococo. En las 526 cepas analizadas se encontraron 42 serotipos; los más frecuentes fueron: 19F (18,1%), 6B (14,3%); 23F (8,9%) y 14 (6,5%). Conclusiones. La distribución de serotipos vacunales en las cepas analizadas fue de 50,0% para los serotipos presentes en la vacuna conjugada heptavalente; 50,2% para los serotipos presentes en la vacuna decavalente y 57,2% para la vacuna 13-valente.

Objectives. To determine the carriage rate and serotype distribution of Streptococcus pneumoniae in the nasopharynx of healthy children younger than 2 years prior to the universal use of the pneumococcal conjugate vaccines in Peru. Materials and methods. Between 2007 and 2009 we collected nasopharyngeal swab samples from 2,123 healthy children aged 2 to 24 months in the vaccination and healthy children consultation offices of pediatric hospitals and health centers in 7 cities in Peru: on the coast (Lima, Piura), highlands (Cusco, Abancay, Arequipa and Huancayo) and amazon basin (Iquitos). The pneumococcal strains were isolated and identified at the central laboratory of the project in Lima, and serotyped by Quellung reaction in the pneumococcal reference laboratory at the Center for Diseases Control and Prevention (CDC). Results. We found 27% (573/2123) of pneumococcal nasopharyngeal healthy carrier children. Among the 526 analyzed strains, we found 42 serotypes; the most common were: 19F (18.1%), 6B (14.3%); 23F (8.9%) and 14 (6.5%). Conclusions. The distribution of vaccine serotypes in the analyzed strains was of 50% for the serotypes present in the seven-valent vaccine, 50.2% for the serotypes present in the ten-valent vaccine and 57.2% for those present in the thirteen-valent vaccine.

[Frequency and pathotypes of diarrheagenic Escherichia coli in Peruvian children with and without diarrhea]. / Frecuencia y patotipos de Escherichia coli diarrogénicas en niños peruanos con y sin diarrea.

UNLABELLED: INTRODUCTION; Diarrheagenic E. coli (DEC) are a major cause of diarrhea in children in developing countries. However, they are not part of routine diagnosis in clinical laboratories. OBJECTIVES: To determine the DEC prevalence in Peruvian children and to describe the genetic variability of these strains. MATERIALS AND METHODS: A total of 8 003 E. coli strains previously isolated from eight different studies of diarrhea in children, mainly from peri-urban areas of Lima, were analyzed. Diagnosis of DEC was done with Multiplex real-time PCR using genes for each of the 6 DEC groups. Conventional PCR was performed for the detection of additional virulence genes. RESULTS: Globally, the mean prevalence in diarrhea samples (n=4,243) was: enteroaggregative E. coli (EAEC) 9.9%, enteropathogenic E. coli (EPEC) 8.5%, enterotoxigenic E. coli (ETEC) 6.9%, diffusely adherent E. coli (DAEC) 4.8%, Shiga toxin-producing E. coli (STEC) 0.8% and enteroinvasive E. coli (EIEC) 0.6%. The relative frequency of each pathogen varies according to the age and the type of study. The main pathotypes in control samples (n=3,760) were EPEC (10.9%) and EAEC (10.4%). An important variability in the virulence genes frequency and molecular resistance mechanisms for each pathotype was found, without differences between diarrhea and control groups. CONCLUSIONS: DEC are a major cause of diarrhea in Peruvian children. These pathogens are highly heterogeneous. Additional studies are required to determine the prevalence in rural areas of Peru and in severe diarrhea cases.

[Adhesion patterns in diffusely adherent Escherichia coli (DAEC) strains isolated from children with and without diarrhea]. / Patrones de adherencia de cepas de Escherichia coli Difusamente adherente (DAEC) provenientes de niños con y sin diarrea.

INTRODUCTION: Diffusely adherent E. coli (DAEC) is the sixth recognized group of diarrheagenic E. coli. However, its association with diarrhea remains controversial. Variability in the adherence patterns of clinical strains is unknown. OBJECTIVES: To compare the adherence patterns between strains isolated from children with and without diarrhea. MATERIALS AND METHODS: A total of 31 DAEC strains were analyzed, 25 from children with diarrhea and 6 from asymptomatic (control) children, isolated from a cohort study of children under one year of age in the southern districts of Lima. DAEC were identified by PCR (daaD gene). The pattern and adherence score in HEp-2 cell culture were evaluated, Actin polimerization was determined by fluorescence actin staining (FAS) and motility was evaluated by conventional microbiology methods. RESULTS: Diffuse adherence pattern was found in 88% of diarrhea samples and in the total of control strains. The number of bacteria adhered per cell was significantly lower in diarrhea samples (p<0.05). However, actin polymerization was greater in diarrhea samples (60% vs. 17%). Motility test was positive in 60% of the diarrhea samples and in all control samples. CONCLUSIONS: Our findings suggest a difference between adherence patterns, actin polymerization and motility between DAEC strains corresponding to diarrhea and control groups. The significance of these results must be confirmed with a bigger number of strains and determining the presence of virulence genes in the strains.

Frecuencia y patotipos de Escherichia coli diarrogénicas en niños peruanos con y sin diarrea / Frequency and pathotypes of diarrheagenic Escherichia coli in peruvian children with and without diarrhea

Introducción. Las E. coli diarrogénicas (DEC) son una de las principales causas de diarrea en niños en países en vías de desarrollo. Sin embargo, no son rutinariamente diagnosticadas en los laboratorios clínicos. Objetivos. Determinar la prevalencia de las DEC en niños peruanos y describir la variabilidad genética de estas cepas. Materiales y métodos. Se utilizaron 8 003 cepas de E. coli previamente aisladas de ocho estudios previos de diarrea en niños, mayormente en zonas periurbanas de Lima. El diagnóstico de las DEC fue a través de un PCR múltiple a tiempo real para los seis grupos de DEC. Se empleó PCR para la determinación de genes adicionales de virulencia. Resultados. La prevalencia promedio global en muestras de diarrea (n=4 243) fue: E. coli enteroagregativa (EAEC) 9,9 por ciento, enteropatogénica (EPEC) 8,5 por ciento, enterotoxigénica (ETEC) 6,9 por ciento, difusamente adherente (DAEC) 4,8 por ciento, productora de toxina shiga (STEC) 0,8 por ciento y enteroinvasiva (EIEC) 0,6 por ciento. La frecuencia relativa de cada patógeno varía según la edad y tipo de estudio. Los principales patotipos en muestras control (n=3 760) fueron EPEC (10,9 por ciento) y EAEC (10,4 por ciento). Se encontró una gran variabilidad en la frecuencia de genes de virulencia para cada patotipo, así como en los mecanismos moleculares de resistencia, sin diferencias significativas entre muestras de diarrea y control. Conclusiones. Las DEC son causa importante de diarrea en niños peruanos. Estos patógenos son altamente heterogéneos. Se requieren estudios adicionales para determinar la prevalencia en zonas rurales del Perú, así como en casos graves de diarrea.

Introduction. Diarrheagenic E. coli (DEC) are a major cause of diarrhea in children in developing countries. However, they are not part of routine diagnosis in clinical laboratories. Objectives. To determine the DEC prevalence in Peruvian children and to describe the genetic variability of these strains. Materials and methods. A total of 8 003 E. coli strains previously isolated from eight different studies of diarrhea in children, mainly from peri-urban areas of Lima, were analyzed. Diagnosis of DEC was done with Multiplex real-time PCR using genes for each of the 6 DEC groups. Conventional PCR was performed for the detection of additional virulence genes. Results. Globally, the mean prevalence in diarrhea samples (n=4,243) was: enteroaggregative E. coli (EAEC) 9.9 percent, enteropathogenic E. coli (EPEC) 8.5 percent, enterotoxigenic E. coli (ETEC) 6.9 percent, diffusely adherent E. coli (DAEC) 4.8 percent, Shiga toxin-producing E. coli (STEC) 0.8 percent and enteroinvasive E. coli (EIEC) 0.6 percent. The relative frequency of each pathogen varies according to the age and the type of study. The main pathotypes in control samples (n=3,760) were EPEC (10.9 percent) and EAEC (10.4 percent). An important variability in the virulence genes frequency and molecular resistance mechanisms for each pathotype was found, without differences between diarrhea and control groups. Conclusions. DEC are a major cause of diarrhea in Peruvian children. These pathogens are highly heterogeneous. Additional studies are required to determine the prevalence in rural areas of Peru and in severe diarrhea cases.

Patrones de adherencia de cepas de Escherichia coli Difusamente adherente (DAEC) provenientes de niños con y sin diarrea / Adhesion patterns in diffusely adherent Escherichia coli (DAEC) strains isolated from children With and without diarrhea

Introducción. Las E. coli de adherencia difusa (DAEC) son el sexto grupo de E. coli diarrogénicas reconocidas. Su asociación con diarrea es controversial. No se conoce la variabilidad en los patrones de adherencia de cepas clínicas. Objetivos. Comparar los patrones de adherencia entre cepas aisladas de niños con y sin diarrea. Materiales y métodos. Se analizó 31 cepas DAEC, 25 de diarrea y 6 de niños asintomáticos (control) aislados de un estudio de cohorte de niños menores de 12 meses en el cono sur de Lima. Las DAEC fueron identificadas por PCR (gen daaD). Se evaluó el patrón y grado de adherencia en cultivos de células HEp-2; la polimerización de actina se evaluó por la prueba de coloración de fluorescencia de actina (FAS); y la motilidad se evaluó por métodos convencionales microbiológicos. Resultados. El patrón de adherencia difusa se encontró en el 88 por ciento de muestras de diarrea y en el 100 por ciento de muestras control. La cantidad de bacterias adheridas por célula fue significativamente menor en las muestras de diarrea (p<0,05). Sin embargo, la polimerización de actina fue mayor en las muestras de diarrea (60 por ciento frente a 17 por ciento). La prueba de motilidad fue positiva en el 60 por ciento de las cepas de diarrea y en el total de muestras control. Conclusiones. Nuestros hallazgos sugieren la existencia de diferencia en los patrones de adherencia, polimerización de actina y motilidad entre cepas de DAEC correspondientes a los grupos de diarrea y control. La significancia de estos resultados debe confirmarse con mayor número de cepas, así como la determinación de los genes de virulencia en las cepas.

Introduction. Diffusely adherent E. coli (DAEC) is the sixth recognized group of diarrheagenic E. coli. However, its association with diarrhea remains controversial. Variability in the adherence patterns of clinical strains is unknown. Objectives. To compare the adherence patterns between strains isolated from children with and without diarrhea. Materials and methods. A total of 31 DAEC strains were analyzed, 25 from children with diarrhea and 6 from asymptomatic (control) children, isolated from a cohort study of children under one year of age in the southern districts of Lima. DAEC were identified by PCR (daaD gene). The pattern and adherence score in HEp-2 cell culture were evaluated, Actin polimerization was determined by fluorescence actin staining (FAS) and motility was evaluated by conventional microbiology methods. Results. Diffuse adherence pattern was found in 88 percent of diarrhea samples and in the total of control strains. The number of bacteria adhered per cell was significantly lower in diarrhea samples (p<0.05). However, actin polymerization was greater in diarrhea samples (60 percent vs. 17 percent). Motility test was positive in 60 percent of the diarrhea samples and in all control samples. Conclusions. Our findings suggest a difference between adherence patterns, actin polymerization and motility between DAEC strains corresponding to diarrhea and control groups. The significance of these results must be confirmed with a bigger number of strains and determining the presence of virulence genes in the strains.

Fecal leukocytes in children infected with diarrheagenic Escherichia coli.

The purpose of this study was to determine the presence and quantity of fecal leukocytes in children infected with diarrheagenic Escherichia coli and to compare these levels between diarrhea and control cases. We analyzed 1,474 stool samples from 935 diarrhea episodes and 539 from healthy controls of a cohort study of children younger than 2 years of age in Lima, Peru. Stools were analyzed for common enteric pathogens, and diarrheagenic E. coli isolates were studied by a multiplex real-time PCR. Stool smears were stained with methylene blue and read by a blinded observer to determine the number of polymorphonuclear leukocytes per high-power field (L/hpf). Fecal leukocytes at >10 L/hpf were present in 11.8% (110/935) of all diarrheal episodes versus 1.1% (6/539) in controls (P < 0.001). Among stool samples with diarrheagenic E. coli as the only pathogen isolated (excluding coinfection), fecal leukocytes at >10 L/hpf were present in 8.5% (18/212) of diarrhea versus 1.3% (2/157) of control samples (P < 0.01). Ninety-five percent of 99 diarrheagenic E. coli diarrhea samples were positive for fecal lactoferrin. Adjusting for the presence of blood in stools, age, sex, undernutrition, and breastfeeding, enterotoxigenic E. coli (ETEC) isolation as a single pathogen, excluding coinfections, was highly associated with the presence of fecal leukocytes (>10 L/hpf) with an odds ratio (OR) of 4.1 (95% confidence interval [CI], 1.08 to 15.51; P < 0.05). Although diarrheagenic E. coli was isolated with similar frequencies in diarrhea and control samples, clearly it was associated with a more inflammatory response during symptomatic infection; however, in general, these pathogens elicited a mild inflammatory response.