Resumo Objetivo Como nosso grupo já havia descrito a composição da microbiota intestinal de neonatos brasileiros em baixo nível socioeconômico, o objetivo deste estudo foi analisar alterações estruturais da comunidade microbiana desse grupo de neonatos no início de sua vida devido a fatores externos. Métodos Amostras fecais foram coletadas mensalmente de 11 neonatos durante o primeiro ano de vida. Os neonatos foram acompanhados com relação a informações clínicas e nutricionais e caracterizados de acordo com práticas de amamentação. O DNA foi extraído das amostras fecais de cada criança e submetido a análise através da técnica de Reação em Cadeia da Polimerase - Eletroforese em Gel de Gradiente Desnaturante. Resultados Os resultados revelaram um padrão de similaridade entre seus próprios pontos temporais em indivíduos em aleitamento materno exclusivo ou predominante. Apesar de variações na intensidade e flutuação de algumas bandas, o padrão Eletroforese em Gel de Gradiente Desnaturante na análise microbiana de um ano foi estável em crianças em aleitamento materno. Houve sucessão ecológica ininterrupta apesar da influência de fatores externos, como alimentação complementar e administração de antibióticos, sugeriu resiliência da microbiota. Isso não foi observado nas crianças com alimentação heterogênea e introdução de alimentos sólidos antes do quinto mês de vida. Conclusão Nossos resultados sugerem um padrão de microbiota intestinal resiliente a forças externas, devido a efeitos probióticos e prebióticos do aleitamento materno exclusivo, reforçam a importância do aleitamento materno exclusivo até o sexto mês de vida.

Abstract Objective Since the present group had already described the composition of the intestinal microbiota of Brazilian infants under low social economic level, the aim of the present study was to analyze the microbial community structure changes in this group of infants during their early life due to external factors. Methods Fecal samples were collected from 11 infants monthly during the first year of life. The infants were followed regarding clinical and diet information and characterized according to breastfeeding practices. DNA was extracted from fecal samples of each child and subjected to Polymerase Chain Reaction - Denaturing Gradient Gel Electrophoresis. Results The results revealed a pattern of similarity between the time points for those who were on exclusive breastfeeding or predominant breastfeeding. Although there were changes in intensity and fluctuation of some bands, the Denaturing Gradient Gel Electrophoresis patterns in the one-year microbial analysis were stable for breastfeeding children. There was uninterrupted ecological succession despite the influence of external factors, such as complementary feeding and antibiotic administration, suggesting microbiota resilience. This was not observed for those children who had mixed feeding and introduction of solid food before the 5th month of life. Conclusion These results suggested an intestinal microbiota pattern resilient to external forces, due to the probiotic and prebiotic effects of exclusive breastfeeding, reinforcing the importance of exclusive breastfeeding until the 6th month of life.

OBJECTIVE: Changes in the neonatal gut environment allow for the colonization of the mucin layer and lumen by anaerobic bacteria. The aim of the present study was to evaluate Bifidobacterium, Lactobacillus and Lactococcus colonization through the first year of life in a group of 12 Brazilian infants and to correlate these data with the levels of Escherichia coli. The presence of anaerobic members of the adult intestinal microbiota, including Eubacterium limosum and Faecalibacterium prausnitzii, was also evaluated. METHODS: Fecal samples were collected during the first year of life, and 16S rRNA from anaerobic and facultative bacteria was detected by real-time PCR. RESULTS: Bifidobacterium was present at the highest levels at all of the studied time points, followed by E. coli and Lactobacillus. E. limosum was rarely detected, and F. prausnitzii was detected only in the samples from the latest time points. CONCLUSION: These results are consistent with reports throughout the world on the community structure of the intestinal microbiota in infants fed a milk diet. Our findings also provide evidence for the influence of the environment on intestinal colonization due to the high abundance of E. coli. The presence of important anaerobic genera was observed in Brazilian infants living at a low socioeconomic level, a result that has already been well established for infants living in developed countries.

OBJECTIVE: The establishment of the intestinal microbiota in newborns is a critical period with possible long-term consequences for human health. In this research, the development of the fecal microbiota of a group of exclusively breastfed neonates living in low socio-economic conditions in the city of São Paulo, Brazil, during the first month of life, was studied. METHODS: Fecal samples were collected from ten neonates on the second, seventh, and 30th days after birth. One of the neonates underwent antibiotic therapy. Molecular techniques were used for analysis; DNA was extracted from the samples, and 16S rRNA libraries were sequenced and phylogenetically analyzed after construction. A real-time polymerase chain reaction (PCR) was performed on the samples taken from the 30th day to amplify DNA from Bifidobacterium sp. RESULTS: The primary phylogenetic groups identified in the samples were Escherichia and Clostridium. Staphylococcus was identified at a low rate. Bifidobacterium sp. was detected in all of the samples collected on the 30th day. In the child who received antibiotics, a reduction in anaerobes and Escherichia, which was associated with an overgrowth of Klebsiella, was observed throughout the experimental period. CONCLUSION: The observed pattern of Escherichia predominance and reduced Staphylococcus colonization is in contrast with the patterns observed in neonates living in developed countries.

No presente estudo, foi encontrado polimorfismo no gene ipa em cinco sorotipos de EIEC, de nove estudados. Quando enzimas de restrição SalI e HindII foram utilizadas no ensaio de PCR-RFLP, amostras de EIEC apresentaram polimorfismo em ipaB e ipaD. Por outro lado, não foram observados polimorfismos nos genes ipaA e ipaC nestas cepas, quando diversas enzimas de restrição foram utilizadas. O polimorfismo presente em cepas de EIEC é sorotipo-dependente, uma vez que os padrões de restrição foram conservados entre as cepas pertencentes ao mesmo sorotipo. Quando a seqüência deduzida de aminoácidos de IpaB de S. flexneri M90T e FBC124-13 foram comparadas, mudanças foram observadas em dez aminoácidos na região amino-terminal. A seqüência deduzida de aminoácidos de IpaD de EIEC apresentou similaridade de 91% com a cepa de Shigella. Neste caso, mudanças de aminoácidos ocorreram em toda a estrutura da molécula de IpaD, exceto na região carboxi-terminal.

In this study, polymorphism in ipa genes was found in five out of nine EIEC serotypes studied. When SalI and HindII were used in RFLP-PCR assays many EIEC serotypes showed polymorphism in ipaB and ipaD. On the other hand, no polymorphism was observed in ipaA and ipaC in these strains. The polymorphism present in EIEC strains is serotype-dependent, since restriction patterns were conserved amongst strains belonging to the same serotype. When IpaB deduced amino acid sequences of S. flexneri M90T and FBC124-13 were compared, ten amino acids changes could be observed mainly in the amino-terminal region. The deduced EIEC IpaD amino-acid sequence presented 91% similarity with the Shigella strain. In this case, amino acid changes were spread out through the whole structure, except in the carboxyl-terminal region.

Objetivo: determinar o perfil etiológico das diarréias agudas de um grupo de crianças de baixo nível socioeconômico atendidas em um serviço regional de pronto-atendimento pediátrico. Método: durante dois anos, as crianças com diarréia aguda atendidas durante um horário pré-estabelecido do dia e da semana foram incluídas no estudo. Os outros critérios seletivos eram: a) idade inferior a 5 anos; b) não utilização de antibiótico no mês precedente; c) ausência de viagem para fora da cidade no mês precedente. Foram pesquisados nas fezes: a) rotavírus (imunofluorescência e contra-imunoeletroforese); b) bactérias - cultura em ágar MacConkey, ágar SS, ágar Columbia, verde brilhante, soroaglutinação, detecção de toxinas - INV, LT,ST,SLT I, SLT II, teste de Séreny, detecção de fatores de virulência -- EAF, eae ,BFP; c) protozoários (Hoffman e Faust). No mesmo período, um grupo controle sem diarréia foi também avaliado para os mesmos patógenos fecais. Resultados: no período de março de 1994 a junho de 1996, foram selecionadas para o estudo 154 crianças com diarréia aguda (GDA) e 42 crianças sem diarréia (GSDA). Foram detectados agentes enteropatogênicos em 112 casos (72,8%) do GDA, e em 9 (21,5%) do GSDA. A associação de dois ou mais enteropatógenos ocorreu em 47 (30,5%) casos do GDA, e em 3 (7,1%) do GSDA. Os patógenos encontrados por caso, do GDA, foram: rotavírus 32 (20,8%), bactérias 53 (34,4%), ambos 25 (16,2%), e 2 (1,4%) com Giardia lamblia (em um caso associada a rotavírus e noutro à bactéria). No GSDA, foram detectadas bactérias em 8 casos (19,1%), e bactéria associada à Giardia lamblia em 1 (2,4%) caso. Das 105 bactérias isoladas no GDA, 90 eram Escherichia coli (EPEC 27, DAEC 24, ETEC 21, EAEC 18), 12 eram Shigella sp, 2 eram Salmonella sp, e uma era Yersinia sp. As crianças com infecção mista - viral e bacteriana - apresentaram maior ocorrência de vômitos repetidos, desidratação e internação.Conclusões: as bactérias foram os enteropatógenos mais detectados nos casos de diarréia aguda, sendo a Escherichia coli a mais freqüente. Na maior parte, as cepas de Escherichia coli eram de biovariedade não-EPEC, habitualmente não investigadas nos laboratórios de patologia clínica. O rotavírus foi encontrado em grande parcela dos casos, muitas vezes em associação com as bactérias. Os protozoários tiveram importância reduzida.

Objective: to evaluate the etiologic profile of acute diarrhea in socioeconomically deprived children assisted at a regional pediatric emergency care service. Methods: during two years all children with acute diarrhea assisted at a previously established day and week time schedule were included in the study. Other selective criteria were: a) age less than 5 years; b) nonuse of antibiotics in the previous month; and c) no travel outside the city in the previous month. Stool examination was used for the detection of the following microorganisms: a) rotavirus (immunofluorescence and counterimmunoelectrophoresis); b) bacteria - culture in MacConkey agar, SS agar, Columbia agar, bright green, serotyping, detection of toxins - INV, LT,ST,SLT I, SLT II, Sereny test, detection of virulence factors - EAF, eae, BFP; and c) protozoa (Hoffman and Faust). In the same period, a control group without diarrhea was also evaluated for the same fecal pathogens. Results: between March 1994 and June 1996, 154 children with acute diarrhea (AD) and 42 control children (WAD), that is, without acute diarrhea, were selected. In the AD group, intestinal pathogens were detected in 112 (72.8%) cases, and in 9 (21.5%) cases in the WAD group. The association of two or more intestinal pathogens occurred in 47 (30.5%) cases in the AD group, and in 3 (7.1%) cases in the WAD group. The pathogens identified in the AD cases were: Rotavirus: 32 (20.8%), bacteria: 53 (34.4%), both: 25 (16.2%), and 2 (1.4%) with Giardia lamblia (in one case associated with Rotavirus and in another one associated with bacteria). In the WAD group, only bacteria were detected in 8 (19.1%) cases, and bacteria associated with Giardia lamblia in 1 (2.4%) case. Altogether, there were 105 bacteria isolated in the AD group: 90 were Escherichia coli (EPEC 27, DAEC 24, ETEC 21, EAEC 18), 12 were Shigella sp, 2 were Salmonella sp, and one was Yersinia sp. Children with mixed infections (viral and bacterial) had increased incidence of severe vomiting, dehydration and hospitalization. Conclusions: bacteria were the most frequent pathogens detected in acute diarrhea cases, among which Escherichia coli was highly predominant. The majority of Escherichia coli strains belong to non-EPEC varieties, strains that are not routinely evaluated in clinical laboratories of pathology. Rotavirus was found in a great number of diarrhea cases, often associated with bacteria. Protozoa showed reduced importance.