Localized, diffuse, and aggregative-adhering Escherichia coli from infants with acute diarrhea and matched-controls

Of 126 infants under 2 years, enrolled in a study on the etiology of acute diarrhea in Recife, Brazil, we selected 37 from whom no recognized enteropathogens, except classic enteropathogenic Escherichia coli, were identified. For comparison, we also examined 37 matched-control infants without diarrhea seen at the same hospital setting. This paper had the purpose to determine the prevalence of localized, diffuse, and aggregative-adhering E. coli strains in both groups. Three to five fecal E. coli colonies, of each case and control, were tested individually for adherence to HeLa cells by using the one step 3-h incubation assay. Strains of E. coli showing localized adherence were found significantly more often in patients (37.8%) than in controls (13.5%), p < 0.02, and they were pratically confined to EPEC serovars 055:H-, 0111:H2, and 119:H6. In contrast, E. coli isolates exhibiting the diffuse or aggregative patterns of adherence were restricted to non-EPEC serogroups and were more frequently encountred among controls.

A Salmonella agona outbreak in a pediatric hospital in the city of Rio de Janeiro, Brazil

Six Salmonella Agona strains from an outbreak of 15 days duration which occurred in a public hospital in Rio de Janeiro, Brazil, were analyzed. The outbreak involved six infants (mean age, 24 days; mean body weight, 1612 g), all of them with severe clinical signs and symptoms. Two of them had surgical implications, two were preterm and two had respiratory distress at birth. The Salmonella strains were resistant to nine antimicrobial agents (ampicillin, cephalotin, cefriaxone, gentamicin, amykacin, trimethoprim-sulfamethoxazole, chloramphenicol, and tetracyclin). Analysis of the plasmid pattern of the wild strains and of the transconjugants confirmed that these were identical strains.

Cryptosporidium sp. in children suffering from acute diarrhea at Uberlândia City, State of Minas Gerais, Brazil

This study's objective was to search for Cryptosporidium sp. in diarrheic feces from children aged zero to 12 years and cared for at medical units within Universidade Federal de Uberlândia or at a private practice in Uberlândia, State of Minas Gerais, Brazil, from September 1992 to August 1993. Three fecal samples preserved in 10% formalin, were collected from 94 children. Oocyst concentration was performed through Ritchie's (modified) method and staining of fecal smears for each sample (total of 1128 slides) was done by the "Safranin/Methylene Blue" and the "Kinyoun (modified)" techniques. The Hoffmann, Pons & Janer method was also employed to look for other enteroparasites. From 94 children, 4.26% excreted fecal Cryptosporidium oocysts. The infection seemed to vary according to age: 5.08% of patients aged zero to two years old; 33.33% of those aging eight to ten years (P>0.05). Cryptosporidium appeared in November, December and March, during the rainy season. 20.21% of the children harbored at least one enteroparasite different from Cryptosporidium, mainly Giardia intestinalis (12.77%). From Cryptosporidium infected patients, two had only this kind, another harbored Giardia intestinalis; the last one hosted Strongyloides stercoralis.

Enteropathogens associated with diarrheal disease in infants of poor urban areas of Porto Velho, Rondônia: a preliminary study

One hundred and thirty cases of diarrhea and 43 age-matched controls, 0 to 5 years old, were studied in a pediatric outpatient unit from a poor peri urban area of Porto Velho, Rondônia. Eighty percent of diarrheal cases were observed in the groups under 2 years of age. Rotavirus (19.2%) was the most frequent enteropathogen associated with diarrhea, followed by Shigella flexneri (6.15%) and S. sonnei (1.5%) and Salmonella sp. (6.9%). Four cases of E. coli enterotoxigenic infections (3.1%), E. coli enteropathogenic (EPEC)(2.3%) one case of E. coli enteroinvasive infection (0.8%) and one case of Yersinia enterocolitica (0.8%) were also identified. Mixed infections were frequent, associating rotavirus, EPEC and Salmonella sp. with Entamoeba histolytica and Giardia lamblia.

Prevalence of Clostridium spp. and Clostridium difficile in children with acute diarrhea in São Paulo city, Brazil

Species of Clostridium are widely distributed in the environment, inhabiting both human and animal gastrointestinal tracts. Clostridium difficile is an important pathogen associated with outbreaks of pseudomembranous colitis and other intestinal disorders, such as diarrhea. In this study, the prevalence of Clostridium spp. and C. difficile, from hospitalized children with acute diarrhea, was examined. These children were admitted to 3 different hospitals for over 12 months. Eighteen (20%) and 19 (21%) stool specimens from children with (90) and without (91) diarrhea respectively, were positive to clostridia. Only 10 C. difficile strains were detected in 5.5% of the stool samples of children with diarrhea. None healthy children (without diarrhea) harbored C. difficile. From these 10 C. difficile, 9 were considered as toxigenic and genotyped as tcdA+/tcdB+ or tcdA-/tcdB+, and 1 strain as nontoxigenic (tcdA-/tdcB-). They were detected by the citotoxicity on VERO cells and by the multiplex-polymerase chain reaction. Thirty clinical fecal extracts produced minor alterations on VERO cells. The presence of C. difficile as a probable agent of acute diarrhea is suggested in several countries, but in this study, the presence of these organisms was not significant. More studies will be necessary to evaluate the role of clostridia or C. difficile in diarrhoeal processes in children.

Detection of cytotoxic necrotizing factor types 1 and 2 among fecal Escherichia coli isolates from brazilian children with and without diarrhea

The enteropathogenic role of cytotoxic necrotizing factor (CNF)-producing Escherichia coli was investigated by searching cnf genes among 2074 isolates from 200 children with and 200 without acute diarrhea in Brazil. Fourteen (7%) cases versus 10 (5%) control children carried at least one cnf positive isolate (P = 0.50) and most isolates expressed CNF type 1. DNA sequences of virulence factors of extraintestinal pathogenic E. coli (ExPEC) were detected in 78.6% of CNF1-producing isolates. Besides not being associated with human acute diarrhea, the CNF1-producing isolates here identified may represent potential ExPEC transitorily composing the normal intestinal flora.

Virulence markers and antimicrobial susceptibility of bacteria of the Bacteroides fragilis group isolated from stool of children with diarrhea in São Paulo, Brazil

Bacteroides fragilis has been isolated from several human and non-human monomicrobial and mixed infections. In this study, some virulence markers and the antimicrobial susceptibility of bacteria of the B. fragilis group isolated from children's stools were evaluated. All the 64 isolates showed the following characteristics: capsulated, beta-hemolytic, hydrophilic, and serum-resistant. Only, 24 (37.5%) strains were resistant at 60ºC, for 30 min, and among them, 12 (18.75%) were resistant at 60ºC, for 60 min. Also, none strain was resistant at 100ºC. Four strains were able to hemagglutinate erythrocytes and D-mannose, D-galactose, D-arabinose, and D-xylose inhibited hemagglutination in 2 B. fragilis strains (p76a, p76b). The hemagglutination in the strain B. uniformis p3-2 was inhibited by D-xylose and D-galactose. The bft gene detection and the enterotoxin production were observed only in 13 EF-enterotoxigenic species. Fragilysin activity was confirmed on HT-29 cells. The antimicrobial determination confirmed that both imipenem and metronidazole were efficient against B. fragilis species; all the strains were resistant to lead and nickel. Plasmids of 2.9, 4.4, 4.8, and 8.9 kb were observed in 6 tested strains. These results show the values of the species identification from clinical infections, as well as of the periodic evaluation of the resistance patterns of the B. fragilis group at Brazilian medical institutions.

Respiratory viruses in the pediatric intensive care unit: prevalence and clinical aspects

A survey was conducted in two pediatric intensive care units in hospitals in Porto Alegre, Brazil, in order to monitor the main respiratory viruses present in bronchiolitis and/or pneumonia and their involvement in the severity of viral respiratory infections. Viral respiratory infection prevalence was 38.7%. In bronchiolitis, respiratory syncytial virus (RSV) was detected in 36% of the cases. In pneumonia, the prevalence rates were similar for adenovirus (10.3%) and RSV (7.7%). There was a difference among the viruses detected in terms of frequency of clinical findings indicating greater severity. Frequency of crackles in patients with RSV (47.3%) showed a borderline significance (p = 0.055, Fisher's exact test) as compared to those with adenovirus (87.5%). The overall case fatality rate in this study was 2.7%, and adenovirus showed a significantly higher case fatality rate (25%) than RSV (2.8%) (p = 0.005). Injected antibiotics were used in 49% of the children with RSV and 60% of those with adenovirus. Adenovirus was not detected in any of the 33 children submitted to oxygen therapy.

Prevalence of diarrheagenic Escherichia coli in children with diarrhea in Salvador, Bahia, Brazil

We report the frequency of the different diarrheagenic Escherichia coli (DEC) categories isolated from children with acute endemic diarrhea in Salvador, Bahia. The E. coli isolates were investigated by colony blot hibridization whit the following genes probes: eae, EAF, bfpA, Stx1, Stx2, ST-Ih, ST-Ip, LT-I, LT-II, INV, and EAEC, as virulence markers to distinguish typical and atypical EPEC, EHEC/STEC, ETEC, EIEC, and EAEC. Seven of the eight categories of DEC were detected. The most frequently isolated was atypical EPEC (10.1%) followed by ETEC (7.5%), and EAEC (4.2%). EHEC, STEC, EIEC, and typical EPEC were each detected once. The strains of ETEC, EAEC, and atypical EPEC belonged to a wide variety of serotypes. The serotypes of the others categories were O26:H11 (EHEC), O21:H21 (STEC), O142:H34 (typical EPEC), and O?H55 (EIEC). We also present the clinical manifestations and other pathogenic species observed in children with DEC. This is the first report of EHEC and STEC in Salvador, and one of the first in Brazil.

Prevalence and genetic diversity of astroviruses in children with and without diarrhea in São Luís, Maranhão, Brazil

Human astroviruses (HAstV) have been increasingly identified as important etiological agents of acute gastroenteritis in children up to five years old. The aim of this study was to determine the prevalence and genotype diversity of HAstV in children with symptomatic and asymptomatic infections in São Luís, Maranhão, Brazil. From June 1997 to July 1999 a total of 183 fecal samples 84 from symptomatic and 99 from asymptomatic children were tested by enzyme immunoassay for HAstV. Prevalence rates were found to be 11 and 3% for symptomatic and asymptomatic children, respectively. Reverse transcription-polymerase chain reaction (RT-PCR) was carried out in 46 specimens (26 symptomatic and 20 asymptomatic) including the 12 samples that were positive by enzyme immunoassay (EIA). The overall positivity yielded by both methods was 8% (15/184); of these, 11% (9/84) for symptomatic and 5% (5/99) for those without symptoms or signs. Sequence analysis of amplicons revealed that HAstV-1 genotype was the most prevalent, accounting for 60% of isolates. Genotypes 2, 3, 4, and 5 were also detected, as one single isolate (10%) for each type. Variations in the sequences were observed when Brazilian isolates were compared to prototype strains identified in the United Kingdom. No seasonal pattern of occurrence was observed during these two years of study, and peak detection rate was observed in children aged between 3 and 6 months in the symptomatic group, and between 18 and 24 months in the controls.

Abnormal humoral immune response to influenza vaccination in pediatric type-1 human immunodeficiency virus infected patients receiving highly active antiretroviral therapy

Given that highly active antiretroviral therapy (HAART) has been demonstrated useful to restore immune competence in type-1 human immunodeficiency virus (HIV-1)-infected subjects, we evaluated the specific antibody response to influenza vaccine in a cohort of HIV-1-infected children on HAART so as to analyze the quality of this immune response in patients under antiretroviral therapy. Sixteen HIV-1-infected children and 10 HIV-1 seronegative controls were immunized with a commercially available trivalent inactivated influenza vaccine containing the strains A/H1N1, A/H3N2, and B. Serum hemagglutinin inhibition (HI) antibody titers were determined for the three viral strains at the time of vaccination and 1 month later. Immunization induced a significantly increased humoral response against the three influenza virus strains in controls, and only against A/H3N2 in HIV-1-infected children. The comparison of post-vaccination HI titers between HIV-1+ patients and HIV-1 negative controls showed significantly higher HI titers against the three strains in controls. In addition, post vaccination protective HI titers (defined as equal to or higher than 1:40) against the strains A/H3N2 and B were observed in a lower proportion of HIV-1+ children than in controls, while a similar proportion of individuals from each group achieved protective HI titers against the A/H1N1 strain. The CD4+ T cell count, CD4/CD8 T cells ratio, and serum viral load were not affected by influenza virus vaccination when pre- vs post-vaccination values were compared. These findings suggest that despite the fact that HAART is efficient in controlling HIV-1 replication and in increasing CD4+ T cell count in HIV-1-infected children, restoration of immune competence and response to cognate antigens remain incomplete, indicating that additional therapeutic strategies are required to achieve a full reconstitution of immune functions.

First detection of a human astrovirus type 8 in a child with diarrhea in Belém, Brazil: comparison with other strains worldwide and identification of possible three lineages

This study describes the genetic relationships of the first human astrovirus type-8 (HAstV-8) detected in Belém-Brazil, during a public hospital-based study. This strain was compared with other HAstV-8 strains identified elsewhere which have sequences available at GeneBank. The regions ORF1a (primers Mon348/Mon340) and ORF2 (primers Mon269/Mon270) were analyzed by nucleotide sequencing and a high similarity rate was observed among the Belém strain and other HAstV-8 strains. In ORF1a, homology values of 93-100% were detected, and in ORF2 96-99%. Considering the sequence variation (7%) observed in ORF2 region, it was suggested that HAstV-8 strains could be divided in three different lineages.

Detection of diarrheagenic Escherichia coli from children with and without diarrhea in Salvador, Bahia, Brazil

We identified different diarrheagenic (DEC) Escherichia coli pathotypes isolated from 1,207 children with and without acute endemic diarrhea in Salvador, Bahia, Brazil collected as part of a case-control study. Since the identification of DEC cannot be based on only biochemical and culture criteria, we used a multiplex polymerase chain reaction developed by combining five specific primer pairs for Enteropathogenic Escherichia coli (EPEC), Shiga toxin-producing E. coli/ Enterohaemorrhagic E. coli (STEC/EHEC), Enterotoxigenic E. coli (ETEC) and Enteroaggregative E. coli (EAEC) to detect these pathotypes simultaneously in a single-step reaction. In order to distinguish typical and atypical EPEC strains, these were tested for the presence of EAF plasmid. The prevalence of diarrheagenic E. coli in this sample of a global case-control study was 25.4% (259 patients) and 18.7% (35 patients) in the diarrhea group (1,020 patients) and the control group (187 patients), respectively. The most frequently isolated pathotype was EAEC (10.7%), followed by atypical EPEC (9.4%), ETEC (3.7%), and STEC (0.6%). Typical EPEC was detected only in one sample. The prevalence of the pathotypes studied in children with diarrhea was not significantly different from that in children without diarrhea.

Hospitalization due to norovirus and genotypes of rotavirus in pediatric patients, state of Espírito Santo

Viruses are the leading cause for hospitalization due to gastroenteritis worldwide. Group A rotaviruses (RV) are the most prevalent and are assorted in glycoproteins (G) and protease sensitive (P) dual genotypes based on polymorphic genes that encode the external VP7 and VP4 capsid proteins, respectively. Noroviruses (NoV) have increasingly answered by sporadic gastroenteritis. This study aimed to determine the prevalence of NoV and RV in 68 hospitalized children, between July 2004 and November 2006, at a pediatric hospital in Vitória city, state of Espírito Santo, Southeastern Brazil. Nucleic acid was extracted from fecal suspension following the guanidine-silica procedure. Reverse transcriptase-polymerase chain reaction (RT-PCR) and polyacrylamide gel electrophoresis were employed for NoV and RV detection, respectively. RV genotyping was accomplished using RT-PCR followed by heminested multiplex PCR with specific primers for the most prevalent types of G and P. Fecal samples were positive for NoV and RV in 39.7% (27/68) and 20.5% (14/68), respectively and together were responsible for 60% (41/68) of the cases. RV genotypes were: 50% G9P[8], 28.7% G2P[4], 7.1% G1P[8], G2P[8] and G?P[8]. Vomit was a prominent manifestation observed in 92% and 85% of the NoV and RV cases, respectively. The median hospitalization was 5 and 5.5 days for the patients infected with NoV and RV, respectively. The data showed that NoV prevailed over RV and it also corroborated the emergence of RV G9 genotype followed by G2P[4], reinforcing the need for RV genotype surveillance.