Infectious etiology and indicators of malabsorption or intestinal injury in childhood diarrhea. / Etiología infecciosa e indicadores de malabsorción o daño intestinal en diarrea infantil

INTRODUCTION: The multifactorial etiology of gastroenteritis emphasizes the need for different laboratory methods to identify or exclude infectious agents and evaluate the severity of diarrheal disease. OBJECTIVE: To diagnose the infectious etiology in diarrheic children and to evaluate some fecal markers associated with intestinal integrity. MATERIALS AND METHODS: The study group comprised 45 children with diarrheal disease, tested for enteropathogens and malabsorption markers, and 76 children whose feces were used for fat evaluation by the traditional and acid steatocrit tests. RESULTS: We observed acute diarrhea in 80% of the children and persistent diarrhea in 20%. Of the diarrheic individuals analyzed, 40% were positive for enteropathogens, with rotavirus (13.3%) and Giardia duodenalis (11.1%) the most frequently diagnosed. Among the infected patients, occult blood was more evident in those carrying pathogenic bacteria (40%) and enteroviruses (40%), while steatorrhea was observed in infections by the protozoa G. duodenalis (35.7%). Children with diarrhea excreted significantly more lipids in feces than non-diarrheic children, as determined by the traditional (p<0.0003) and acid steatocrit (p<0.0001) methods. Moreover, the acid steatocrit method detected 16.7% more fecal fat than the traditional method. CONCLUSIONS: Childhood diarrhea can lead to increasingly severe nutrient deficiencies. Steatorrhea is the hallmark of malabsorption, and a stool test, such as the acid steatocrit, can be routinely used as a laboratory tool for the semi-quantitative evaluation of fat malabsorption in diarrheic children.

Introducción. La etiología multifactorial de la gastroenteritis enfatiza la necesidad de usar diferentes métodos de laboratorio para identificar o excluir agentes infecciosos y evaluar la gravedad de la enfermedad diarreica. Objetivo. Diagnosticar la etiología infecciosa de la diarrea en niños y evaluar algunos marcadores fecales asociados con la integridad intestinal. Materiales y métodos. Se estudiaron 45 niños con enfermedad diarreica, en los cuales se evaluaron la presencia de enteropatógenos y los marcadores de malabsorción. Se analizaron las muestras fecales de 76 niños, mediante las pruebas de esteatocrito tradicional y esteatocrito ácido, para la cuantificación de la grasa. Resultados. Se observó diarrea aguda en el 80 % de los niños y diarrea persistente en el 20 %. De los individuos con diarrea, el 40 % fue positivo para enteropatógenos; los más diagnosticados fueron rotavirus (13,3 %) y Giardia duodenalis (11,1 %). Entre los pacientes infectados, la sangre oculta fue más evidente en aquellos portadores de bacterias patógenas (40 %) o enterovirus (40%), mientras que la esteatorrea se observó en infecciones por el protozoo G. duodenalis (35,7 %). Los niños con diarrea excretaron significativamente más lípidos en las heces que aquellos sin diarrea, según lo determinado por los métodos de esteatocrito tradicional (p<0,0003) y esteatocrito ácido (p<0,0001). Conclusiones. La diarrea infantil puede provocar deficiencias graves de nutrientes. La esteatorrea es distintiva de la malabsorción intestinal y puede detectarse mediante la estimación del esteatocrito ácido. Esta prueba podría utilizarse de forma rutinaria como una herramienta de laboratorio para la evaluación semicuantitativa de la malabsorción de grasas en niños con diarrea.

Cryptosporidium diagnosis in different groups of children and characterization of parasite species.

BACKGROUND: Microscopy and enzyme-linked immunosorbent assay (ELISA) are routinely used for Cryptosporidium diagnosis, without differentiating the parasite species. METHODS: Children's feces were analyzed by modified Ziehl-Neelsen (mZN) and ELISA for Cryptosporidium diagnosis and by polymerase chain reaction-restriction fragment length polymorphism for species identification. RESULTS: Cryptosporidium frequency was 2.6%. The sensitivity and specificity of ELISA were 85.7% and 99.7%, respectively, with excellent concordance with mZN (kappa=0.854). Parasite species were characterized as Cryptosporidium hominis (78.3%), Cryptosporidium felis (17.4%), and Cryptosporidium parvum (4.3%). CONCLUSIONS: Coproantigen ELISA is as efficient as mZN for Cryptosporidium diagnosis. Cryptosporidium genotyping suggests anthroponotic and zoonotic transmission to children.

Cryptosporidium diagnosis in different groups of children and characterization of parasite species

ABSTRACT Background: Microscopy and enzyme-linked immunosorbent assay (ELISA) are routinely used for Cryptosporidium diagnosis, without differentiating the parasite species. Methods: Children's feces were analyzed by modified Ziehl-Neelsen (mZN) and ELISA for Cryptosporidium diagnosis and by polymerase chain reaction-restriction fragment length polymorphism for species identification. Results: Cryptosporidium frequency was 2.6%. The sensitivity and specificity of ELISA were 85.7% and 99.7%, respectively, with excellent concordance with mZN (kappa=0.854). Parasite species were characterized as Cryptosporidium hominis (78.3%), Cryptosporidium felis (17.4%), and Cryptosporidium parvum (4.3%). Conclusions: Coproantigen ELISA is as efficient as mZN for Cryptosporidium diagnosis. Cryptosporidium genotyping suggests anthroponotic and zoonotic transmission to children.

The Predominance of Giardia duodenalis AII sub-assemblage in young children from Salvador, Bahia, Brazil. / Predominio del subconjunto AII de Giardia duodenalis en niños pequeños de Salvador, Bahía, Brasil

INTRODUCTION: Giardia duodenalis is an intestinal protozoan with a high prevalence in children of developing countries. Molecular studies revealed a great genetic diversity of G. duodenalis, with assemblages A and B found mainly in humans. Despite its importance, the information on the molecular epidemiology of human giardiasis is still limited in Brazil. OBJECTIVE: To characterize G. duodenalis molecular isolates in children from Salvador, Bahia, Brazil. MATERIALS AND METHODS: Giardia duodenalis positive fecal samples were obtained from 71 children from two day care centers and 39 users of a clinical analysis laboratory. Samples were analyzed by PCR-RFLP of the glutamate dehydrogenase (gdh) and beta-giardin genes and by the sequencing of beta-giardin. RESULTS: Of the 110 G. duodenalis samples, 80 (72.7%) amplified one or both target genes. Of these, 62 (77.5 %) were identified as assemblage A and 18 (22.5%) as assemblage B. The subassemblage AII was identified in 58.8% (n=47) of isolates followed by the sub-assemblage AI (18.8%, n=15), BIV (11.2%, n=9), and BIII (5.0%, n=4). The AII sub-assemblage was the most frequent in children of both day care centers whereas AI was found only in the group attended at the clinical laboratory. Sub-assemblage AII predominated in children under two years. CONCLUSIONS: The higher frequency of AII sub-assemblage suggests that anthroponotic transmission is more common in Salvador, but that zoonotic transmission pathways are also present and a change in susceptibility to different molecular patterns of Giardia may occur during child growth.

Introducción. Giardia duodenalis es un protozoo intestinal de gran prevalencia en los niños de los países en desarrollo. En estudios moleculares se ha evidenciado la gran diversidad genética de G. duodenalis y se han identificado los conjuntos A y B, principalmente en humanos. A pesar de su importancia, el conocimiento de la epidemiología molecular de la giardiasis humana aún es limitado en Brasil. Objetivo. Caracterizar los aislamientos moleculares de G. duodenalis de muestras tomadas a niños de Salvador, Bahía, Brasil. Materiales y métodos. Las muestras fecales positivas para G. duodenalis se obtuvieron de 71 niños de dos guarderías y de 39 usuarios de un laboratorio de análisis clínicos. Las muestras se analizaron mediante PCR-RFLP de los genes gdh y beta-giardin, y secuenciación de beta-giardin. Resultados. De las 110 muestras de G. duodenalis, en 80 (72,7 %) se amplificaron uno o ambos genes. De estos, 62 (77,5 %) se identificaron como pertenecientes al conjunto A y 18 (22,5 %) al B. El subconjunto AII se identificó en el 58,8 % (n=47) de los aislamientos, seguido del AI en el 18,8% (n=15), el BIV en el 11,2% (n=9) y el BIII en el 5,0% (n=4). El subconjunto AII fue el más frecuente en los niños de ambas guarderías, en tanto que el AI solo se encontró en el grupo atendido en el laboratorio clínico. El subconjunto AII predominó en los niños menores de dos años. Conclusiones. La mayor frecuencia del subconjunto AII sugiere que la transmisión antroponótica es más común en Salvador, pero también existen vías de transmisión zoonóticas, y que pueden ocurrir cambios en la sensibilidad frente a diferentes patrones moleculares de Giardia durante el crecimiento infantil.

Predominance of Giardia duodenalis AII sub-assemblage in young children from Salvador, Bahia, Brazil / Predominio del subconjunto AII de Giardia duodenalis en niños pequeños de Salvador, Bahía, Brasil

Introduction. Giardia duodenalis is an intestinal protozoan with a high prevalence in children of developing countries. Molecular studies revealed a great genetic diversity of G. duodenalis, with assemblages A and B found mainly in humans. Despite its importance, the information on the molecular epidemiology of human giardiasis is still limited in Brazil. Objective. To characterize G. duodenalis molecular isolates in children from Salvador, Bahia, Brazil. Materials and methods. Giardia duodenalis positive fecal samples were obtained from 71 children from two day care centers and 39 users of a clinical analysis laboratory. Samples were analyzed by PCR-RFLP of the glutamate dehydrogenase (gdh) and beta-giardin genes and by the sequencing of beta-giardin. Results. Of the 110 G. duodenalis samples, 80 (72.7%) amplified one or both target genes. Of these, 62 (77.5 %) were identified as assemblage A and 18 (22.5%) as assemblage B. The subassemblage AII was identified in 58.8% (n=47) of isolates followed by the sub-assemblage AI (18.8%, n=15), BIV (11.2%, n=9), and BIII (5.0%, n=4). The AII sub-assemblage was the most frequent in children of both day care centers whereas AI was found only in the group attended at the clinical laboratory. Sub-assemblage AII predominated in children under two years. Conclusions. The higher frequency of AII sub-assemblage suggests that anthroponotic transmission is more common in Salvador, but that zoonotic transmission pathways are also present and a change in susceptibility to different molecular patterns of Giardia may occur during child growth.

Introducción. Giardia duodenalis es un protozoo intestinal de gran prevalencia en los niños de los países en desarrollo. En estudios moleculares se ha evidenciado la gran diversidad genética de G. duodenalis y se han identificado los conjuntos A y B, principalmente en humanos. A pesar de su importancia, el conocimiento de la epidemiología molecular de la giardiasis humana aún es limitado en Brasil. Objetivo. Caracterizar los aislamientos moleculares de G. duodenalis de muestras tomadas a niños de Salvador, Bahía, Brasil. Materiales y métodos. Las muestras fecales positivas para G. duodenalis se obtuvieron de 71 niños de dos guarderías y de 39 usuarios de un laboratorio de análisis clínicos. Las muestras se analizaron mediante PCR-RFLP de los genes gdh y beta-giardin, y secuenciación de beta-giardin. Resultados. De las 110 muestras de G. duodenalis, en 80 (72,7 %) se amplificaron uno o ambos genes. De estos, 62 (77,5 %) se identificaron como pertenecientes al conjunto A y 18 (22,5 %) al B. El subconjunto AII se identificó en el 58,8 % (n=47) de los aislamientos, seguido del AI en el 18,8% (n=15), el BIV en el 11,2% (n=9) y el BIII en el 5,0% (n=4). El subconjunto AII fue el más frecuente en los niños de ambas guarderías, en tanto que el AI solo se encontró en el grupo atendido en el laboratorio clínico. El subconjunto AII predominó en los niños menores de dos años. Conclusiones. La mayor frecuencia del subconjunto AII sugiere que la transmisión antroponótica es más común en Salvador, pero también existen vías de transmisión zoonóticas, y que pueden ocurrir cambios en la sensibilidad frente a diferentes patrones moleculares de Giardia durante el crecimiento infantil.

Specific IgG and IgA Antibody Reactivities in Sera of Children by Enzyme-Linked Immunoassay and Comparison With Giardia duodenalis Diagnosis in Feces.

BACKGROUND: Giardia duodenalis is conventionally diagnosed in fecal samples using parasitological methods. However, sensitivity is poor when only a single sample is analyzed, due to intermittent excretion of cysts in feces. Alternatively, the serum antibodies to G. duodenalis can be used for parasite diagnosis and epidemiological studies to determine previous exposure. We compared the rate of G. duodenalis infection between serum anti-Giardia IgG and IgA antibodies and fecal examination in Brazilian children. METHODS: Fecal and serum samples were tested from 287 children at a clinical laboratory and from 187 children at daycare centers. Fecal samples were processed using conventional parasitological methods and coproantigen detection for Giardia diagnosis. Serum samples were tested using an in-house ELISA for detection of anti-Giardia IgG and IgA. RESULTS: G. duodenalis was found in 8.2% (N=39) of the 474 children analyzed. The sensitivity and specificity of ELISA were 80.0% and 90.0% for IgG and 80.0% and 83.3% for IgA, respectively. The total positivity rate of anti-Giardia IgG and IgA in the sera was 13.9% (N=66) and 23.6% (N=112). The agreement between the positivity of specific antibodies and the detection of G. duodenalis in feces was moderate for ELISA-IgG, kappa index (95% CI)=0.543 (0.422-0.664), and mild for ELISA-IgA, kappa index (95% CI)=0.283 (0.162-0.404). Among the children infected with other enteroparasites, 11.6% (N=10) and 24.4% (N=21) showed reactivity to anti-Giardia IgG and to IgA, respectively. This cross-reactivity was more frequent in samples from children infected with Endolimax nana and Entamoeba coli. CONCLUSIONS: The higher frequency of specific antibody reactivity compared with G. duodenalis diagnosis in feces could reflect continuous exposure of children to G. duodenalis infection, resulting in long-lasting immunological memory and/or cross-reactivity with other intestinal amoebas.

Milk protein-based formulas containing different oils affect fatty acids balance in term infants: A randomized blinded crossover clinical trial.

BACKGROUND: Palm olein is used in infant formula fat blends in order to match the fatty acid profile of human milk. While the effects on fatty acid balance have been evaluated, the use of palm olein in combination with palm kernel oil and supplementation with docosahexaenoic acid (DHA) and arachidonic acid (ARA) has not been similarly assessed in infants. This study evaluated the effects of infant formulas containing different fat compositions on the balance of fat, fatty acids, and calcium. METHODS: In this randomized, crossover, double-blinded study, 33 healthy term infants (68-159 ± 3 days of age at enrollment) were fed two formulas for 14 days in a tolerance period, followed by a 4-day metabolic balance period in 17 of the male subjects. The study compared two commercially available milk-based powdered formulas in Brazil; the PALM formula contained palm olein (44%), kernel palm oil (21.7%), and canola oil (18.5%) as the predominant fats, whereas the NoPALM formula contained other fat sources. RESULTS: Fat absorption (%) was greater for NoPALM versus PALM-fed infants (96.55 and 95.50%, respectively; p = 0.023). The absorption percentage of palmitic acid (C16:0) did not differ significantly between formulas (p > 0.05), but this acid was excreted at significantly higher concentrations in the PALM (29.42 mg/kg/day) than in the NoPALM (12.28 mg/kg/day) formula groups. DHA and ARA absorption percentages were also higher in NoPALM-fed infants. Calcium absorption was higher in NoPALM-fed infants (58.00%) compared to those fed PALM (40.90%), but the difference was not significant (p = 0.104) when calcium intake was used as a covariate. However, calcium retention was higher in NoPALM-fed infants compared to that in PALM-fed infants with or without calcium intake as a covariate. Adverse events did not differ between groups (p > 0.05). CONCLUSIONS: The absorption of essential fatty acids was similar for both formulas; however, long-chain polyunsaturated fatty acids (DHA and ARA) were better absorbed from the NoPALM formula. Fat absorption and calcium retention were lower in term infants fed the PALM-based formula. CLINICAL TRIAL REGISTRATION: Clinicaltrial.gov # NCT00941564 .

Normal Growth of Healthy Infants Born from HIV+ Mothers Fed a Reduced Protein Infant Formula Containing the Prebiotics Galacto-Oligosaccharides and Fructo-Oligosaccharides: A Randomized Controlled Trial.

OBJECTIVE: The aim of the current study was to evaluate the safety of a new reduced protein (2.1 g/100 kcal) infant formula containing 4 g/L of 90% galacto-oligosaccharides (GOS) and 10% fructo-oligosaccharides (FOS). METHODS: Healthy term infants from Brazil were enrolled. Those born to human immunodeficiency virus (HIV)-positive mothers were randomized to a test (n = 65) or control (n = 63) formula group. Infants born to HIV-negative mothers were either exclusively breast-fed (n = 79) or received a mixed diet (breast milk and test formula, n = 65). Between 2 weeks and 4 months of age, infants were exclusively fed according to their assigned group. Anthropometric measurements were taken at baseline, 1, 2, 3, 4, 6, 8, 10, and 12 months. Digestive tolerance was evaluated during the first 4 months. The primary outcome was mean daily weight gain between 2 weeks and 4 months in the test formula and breast-fed groups. RESULTS: Data from all infants (N = 272) were used in the intention-to-treat (ITT) analysis and data from 230 infants were used in the per-protocol (PP) analysis. The difference in mean daily weight gain between 2 weeks and 4 months in the test formula and breast-fed groups was 1.257 g/day (one-sided 95% confidence interval [CI]: -0.705 to inf, P < 0.001) in the PP analysis, showing that the lower bound of the 95% CI was above the -3.0 g/day non-inferiority margin. Results were similar in the ITT analysis. Symptoms of digestive tolerance and frequency of adverse events were similar in the two groups. CONCLUSIONS: The formula containing 2.1 g/100 kcal protein and GOS and FOS was safe and tolerated well.

Segmental absence of intestinal musculature -presentation in a 10-year-old boy with an extensive involved segment.

Segmental absence of the intestinal musculature is a rare cause of acute abdomen most likely associated with intestinal perforation and sepsis in neonates and adults. We present a case of a 10-year-old boy who developed acute abdomen and was treated with right hemicolectomy and partial resection of the ileum. The ileum showed a 20-cm-long stenotic segment showing prominent thinning of the intestinal wall. This case occurred in an older child, which appears to be rare when comparing the literature. The bowel defect was also the largest described to date, indicating the potential effect of an atonic segment leading to obstructive disease.

Suplementação de zinco e outros micronutrientes através do uso de sprinkles: impacto na ocorrência de doença diarreica e infecções respiratórias em crianças institucionalizadas / Zinc and other micronutrients supplementation through the use of sprinkles: impact on the occurrence of diarrhea and respiratory infections in institutionalized children

OBJETIVO: Avaliar a incidência de doenças diarreicas (DA) e infecção respiratória aguda (IRA) em crianças submetidas à suplementação de zinco e outros micronutrientes através dos sprinkles, bem como a aceitação destes pelos participantes. MÉTODO: Ensaio clínico, duplo cego, randomizado, realizado com 143 crianças institucionalizadas, saudáveis, de seis a 48 meses. As mesmas foram randomizadas em dois grupos e receberam diariamente zinco + micronutrientes - grupo teste (sprinkles), ou apenas micronutrientes sem zinco - grupo controle. As crianças foram suplementadas por 90 dias e acompanhadas quanto aos desfechos de DA e IRA. RESULTADOS: Das crianças randomizadas, 52,45% pertenciam ao grupo teste e 47,55% ao controle. A incidência de DA no teste foi de 14,7%, e no controle, 19,1%. O grupo teste apresentou menor risco de desenvolver DA em relação ao controle, porém esse achado não foi estatisticamente significante (RR = 0,77 [0,37-1,6]; p = 0,5088). A IRA apresentou incidência elevada em ambos os grupos, sendo 60% no teste e 48,5% no controle, com risco maior de apresentar a doença no grupo teste, porém sem significância estatística (RR=1,24 [0,91-1,68]; p = 0,1825). Quanto à aceitação, o percentual médio de consumo, em dias, de todo conteúdo dos sachês contendo sprinkles foi 95,72% (DP = 4,9) e 96,4% (DP = 6,2), para o teste e controle, respectivamente. CONCLUSÕES: A suplementação de zinco através dos sprinkles não reduziu a incidência de DA ou IRA entre as crianças avaliadas. Os sprinkles foram bem aceitos por todos os participantes do estudo.

OBJECTIVE: To evaluate the incidence of diarrheal disease (DD) and acute respiratory infection (ARI) in children undergoing supplementation of zinc and other micronutrients through the use of sprinkles, as well as their acceptance by these participants. METHOD: This was a randomized double-blinded clinical trial of 143 healthy institutionalized children, aged 6 to 48 months. They were randomized into two groups and received daily zinc and micronutrients - test group (sprinkles), or micronutrients without zinc - control group. Children were supplemented for 90 days and followed regarding the outcomes of DD and ARI. RESULTS: Of the randomized children, 52.45% belonged to the test and 47.55% to the control group. The incidence of DD in the test group was 14.7% and was 19.1% in the control group. The test group showed a lower risk of developing DD when compared to controls, but this finding was not statistically significant (RR = 0.77 [0.37 to 1.6], p = 0.5088). ARI had high incidence in both groups, 60% in the test group and 48.5% in the control group, with an increased risk of developing the disease in the test group, but with no statistical significance (RR = 1.24 [0.91 to 1.68], p = 0.1825). Regarding acceptance, the mean percentage of consumption, in days, of the entire content of the sachets containing sprinkles was 95.72% (SD = 4.9) and 96.4% (SD = 6.2) for the test and control groups, respectively. CONCLUSIONS: Zinc supplementation through the use of sprinkles did not reduce the incidence of DD or ARI among the evaluated children. The sprinkles were well accepted by all study participants.

Zinc and other micronutrients supplementation through the use of sprinkles: impact on the occurrence of diarrhea and respiratory infections in institutionalized children.

OBJECTIVE: To evaluate the incidence of diarrheal disease (DD) and acute respiratory infection (ARI) in children undergoing supplementation of zinc and other micronutrients through the use of sprinkles, as well as their acceptance by these patients. METHOD: This was a randomized double-blinded clinical trial of 143 healthy institutionalized children, aged 6 to 48 months. They were randomized into two groups and received daily zinc and micronutrients - test group (sprinkles), or micronutrients without zinc - control group. Children were supplemented for 90 days and followed regarding the outcomes of DD and ARI. RESULTS: Of the randomized children, 52.45% belonged to the test and 47.55% to the control group. The incidence of DD in the test group was 14.7% and was 19.1% in the control group. The test group showed a lower risk of developing DD when compared to controls, but this finding was not statistically significant (RR=0.77 [0.37 to 1.6], p=0.5088). ARI had high incidence in both groups, 60% in the test group and 48.5% in the control group, with an increased risk of developing the disease in the test group, but with no statistical significance (RR=1.24 [0.91 to 1.68], p=0.1825). Regarding acceptance, the mean percentage of consumption, in days, of the entire content of the sachets containing sprinkles was 95.72% (SD=4.9) and 96.4% (SD=6.2) for the test and control groups, respectively. CONCLUSIONS: Zinc supplementation through the use of sprinkles did not reduce the incidence of DD or ARI among the evaluated children. The sprinkles were well accepted by all study participants.

Cosavirus infection in persons with and without gastroenteritis, Brazil.

To determine possible cosavirus association with clinical disease, we used real-time reverse transcription PCR to test children and HIV-positive adults in Brazil with and without gastroenteritis. Thirteen (3.6%) of 359 children with gastroenteritis tested positive, as did 69 (33.8%) of 204 controls. Low prevalence, frequent viral co-infections, and low fecal cosavirus RNA concentrations argue against human pathogenicity.

Full genome sequence analysis of parechoviruses from Brazil reveals geographical patterns in the evolution of non-structural genes and intratypic recombination in the capsid region.

Due to high genome plasticity, the evolutionary fate and geographical history of picornaviruses is hard to follow. Here, we determined the complete coding sequences of eight human parechoviruses (HPeV) of types 1, 5 and 6 directly from clinical samples from Brazil. The capsid genes of these strains were not remarkably different from European, North American and Japanese HPeV. Full genome analysis revealed frequent intertypic recombination in the non-structural genome region. In addition, evidence of recombination between viruses of the same type in the capsid-encoding genome region among HPeV1 and HPeV4 was obtained. Bayesian phylogenetic analysis indicated that strains without evidence of recombination with each other in any genome region were separated by no more than 35 years of circulation. Interestingly, in the 3C gene, all Brazilian parechoviruses grouped together regardless of serotype. The most recent common ancestor of these strains dated back 108 years, suggesting long-term endemicity of this particular P3 genome lineage in South America. Our results support the idea that picornavirus replicative genes acquire capsid proteins introduced by new strains. Under certain epidemiological conditions, replicative genes may be maintained in circumscript geographical regions.

Genomic features and evolutionary constraints in Saffold-like cardioviruses.

This study identified the complete genomic sequence of four type 2 and type 3 human Saffold-like cardioviruses (SLCVs) isolated in Germany and Brazil. The secondary structures of the SLCV internal ribosome entry sites (IRESs) were deduced based on RNA base-pairing conservation and co-variation, using an established Theiler's murine encephalomyelitis virus (TMEV) IRES structure as a reference. The SLCV IRES was highly similar to that of TMEV, but motifs critical in TMEV for binding of the polypyrimidine tract-binding protein (PTB) were disrupted. In TMEV, corresponding alterations have been associated with reduced neurovirulence in mice. In the non-structural genome region, there was evidence of multiple intertypic recombination events between different SLCV types. Between viruses of the same type, recombination also occurred in the capsid-encoding genome region. There were apparently no recombination events between mouse TMEV and human SLCV. In another genus of the family Picornaviridae, Enterovirus, natural recombination occurs strictly within species and can serve as an additional criterion for delimiting species. Accordingly, the results of this study suggest that SLCV and TMEV may represent distinct species within the genus Cardiovirus.

Lack of effect of nucleotide-supplemented infant formula on the management of acute diarrhea in infants.

Nucleotides have been identified as conditionally essential nutrients. As prevention studies, conducted with nucleotide-supplemented formula, have shown statistically significant decrease in the risk of diarrhea, we tested the hypothesis that the consumption of nucleotide-supplemented formula during an acute diarrhea episode is associated with therapeutic effects in the treatment of infants with acute diarrhea and dehydration. A randomized, double-blind, controlled clinical trial was conducted in which patients were randomly assigned to 1 of 2 treatment groups. The "test" group consumed a nucleotide-supplemented infant formula and the "control" group consumed a nonsupplemented formula. Infants were accommodated in a metabolic unit where body weight, and all intakes and outputs were recorded at 24-hour intervals during hospitalization. Laboratory parameters including blood gases and electrolytes were monitored during hospitalization. Eighty-one male infants ranging in age from older than 1 month and younger than 1 year, with acute non-cholera diarrhea and dehydration were studied. Primary outcomes were stool output and duration of diarrhea and did not differ significantly between the groups, with a stool output of 304.2 (SD 254.0) vs 350.3 (SD 269.1) g/kg and a duration of diarrhea of 83.3 (SD 44.5) vs 88.8 (SD 46.6) for the test and control groups, respectively. Anemia was highly prevalent and breast-feeding practice was not frequent in both groups. The average energy intake and weight gain were similar in the 2 groups. This study demonstrated that nucleotide supplementation of infant formula during episodes of acute diarrhea has no therapeutic advantage compared to conventional infant formula.

Novel human parechovirus from Brazil.

Human parechoviruses (HPeVs) were detected by reverse transcription-PCR in 16.1% of 335 stool samples from children <6 years of age with enteritis in Salvador, Brazil. Whole genome sequencing of 1 sample showed a novel HPeV that has been designated as HPeV8.

Circulation of 3 lineages of a novel Saffold cardiovirus in humans.

Cardioviruses cause serious disease, mainly in rodents, including diabetes, myocarditis, encephalomyelitis, and multiple sclerosis-like disseminated encephalomyelitis. Recently, a human virus isolate obtained 25 years ago, termed Saffold virus, was sequenced and classified as a cardiovirus. We conducted systematic molecular screening for Saffold-like viruses in 844 fecal samples from patients with gastroenteritis from Germany and Brazil, across all age groups. Six cardioviruses were identified in patients <6 years of age. Viral loads were 283,305-5,044,412,175 copies/g of stool. Co-infections occurred in 4 of 6 children. No evidence for outbreak-like epidemic patterns was found. Phylogenetic analysis identified 3 distinct genetic lineages. Viral protein 1 amino acids were 67.9%-77.7% identical and had a distance of at least 39.4% from known cardioviruses. Because closely related strains were found on 2 continents, global distribution in humans is suspected. Saffold-like viruses may be the first human cardiovirus species to be identified.

Controlled, double-blind, randomized clinical trial to evaluate the impact of fruit juice consumption on the evolution of infants with acute diarrhea.

In order to assess the effects of juice feedings during acute diarrhea a double-blind, randomized study was performed in 90 children, mean age of 10 +/- 4.28 months. Thirty patients with acute diarrhea were fed twice-daily 15 ml/kg of Apple Juice (AJ), 30 received White Grape Juice (WGJ), and 30 were given colored and flavored water (WA) as part of their age appropriate dietary intake. The duration and severity of diarrhea were the main endpoint variables of the study performed in a metabolic unit. The patients were similar among the 3 groups, had diarrhea for 50-64 hours prior to admission, and were dehydrated when admitted to the unit for study. Half of the patients in each group were well nourished and the others had mild to moderate degrees of malnutrition. Rotavirus infection was the agent causing the illness in 63% of the patients. The infants fed juice ingested 14-17% more calories than those given WA, (those receiving AJ and WGJ ingested 95 and 98 Calories/Kg/d respectively) whereas those receiving WA consumed 81 cal/kg/d). The increased energy intake was not at the expense of other foods or milk formula. The mean body weight gain was greater among patients receiving WGJ (+ 50.7 gm) as compared with the patients in the AJ group (+ 18.3 gm) or the patients fed WA (- 0.7 gm) (p = 0.08). The duration of the illness was longer in the infants fed juice as compared with those given WA (p = 0.006), the mean +/- SD duration in hours was 49.4 +/- 32.6, 47.5 +/- 38.9 and 26.5 +/- 27.4 in patients fed AJ, WGJ and WA respectively. All patients improved while ingesting juice and none of them developed persistent diarrhea; most recovered within 50 hours of the beginning of treatment and less than one fourth had diarrhea longer than 96 hours in the unit. The fecal losses were also increased among the juice fed patients (p = 0.001); the mean +/- SD fecal excretion in g/kg/h was 3.94 +/- 2.35, 3.59 +/- 2.35, and 2.19 +/- 1.63 in AJ, WGJ and WA respectively. The stool output was highest during the first day of treatment among all the patients, though those fed AJ had the highest volume of fecal losses and those who received WA had the lowest stool excretion. After the first day of treatment the differences in fecal excretion were not significant. The ability to tolerate carbohydrates during the illness and immediately after recovery was similar among the 3 groups of patients. Intake of juices with different fructose/glucose ratios and osmolarities resulted in more fecal losses and more prolonged diarrhea as compared with water feedings, but the patients given juice ingested more calories and gained more weight, particularly among those being fed the juice with equimolar concentrations of fructose and glucose.

Limitations of probiotic therapy in acute, severe dehydrating diarrhea.

BACKGROUND: Recent studies have shown that probiotics, most commonly, may be useful in treating acute gastroenteritis. However, beneficial effects appear to be limited to a modest decrease in the duration of diarrhea. No studies have evaluated this therapy in moderate to severe dehydrating diarrhea in a metabolic facility. METHODS: Male children less than 2 years of age were admitted to a metabolic unit of the Department of Pediatrics at the Federal University of Bahia, Brazil, with moderate dehydration and were randomized in a double-blind, placebo-controlled fashion. Oral rehydration solution (ORS) was administered per protocol and either placebo or was given in combination with the ORS. Output of urine, stool, and vomitus was recorded along with stool weight, nude body weight, and standard laboratory assessments for hydration. RESULTS: There was no significant reduction in diarrhea duration and stool output in the group. However, Kaplan-Meier survival analysis demonstrated that, even in moderate to severe diarrhea, resolution of the illness occurred so rapidly, that statistically significant benefits of probiotic therapy could not be demonstrated. CONCLUSION: Our data implies that colonization must occur before benefits of probiotics can be realized. Probiotics are, therefore, likely to be of limited benefit in treating diarrheal illnesses of short duration such as viral enteritis. The beneficial effects of probiotics may be limited to prophylactic usage in high-risk populations.